1-thia-2,4a-diaza-cyclopenta[b]napththalene-3,4-diones and related compounds as anti-infective agents

ABSTRACT

The invention provides compounds and salts of Formula I and Formula II: 
                         
which possess antimicrobial activity. The invention also provides novel synthetic intermediates useful in making compounds of Formula I and Formula II. The variables A 1 , A 8 , R 2 , R 3 , R 5 , R 6 , R 7 , and R 9  are defined herein.
 
     Certain compounds of Formula I and Formula II disclosed herein are potent and selective inhibitors of bacterial DNA synthesis and bacterial replication. The invention also provides antimicrobial compositions, including pharmaceutical compositions, containing one or more compounds of Formula I or Formula II and one or more carriers, excipients, or diluents. Such compositions may contain a compound of Formula I or Formula II as the only active agent or may contain a combination of a compound of Formula I or Formula II and one or more other active agents. The invention also provides methods for treating microbial infections in animals.

CROSS REFERENCE TO RELATED APPLICATION

This application claims priority from U.S. Provisional application60/641,548 filed Jan. 5, 2005, which is hereby incorporated by referencein its entirety.

FIELD OF THE INVENTION

The present invention provides1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-diones and relatedcompounds, which possess antimicrobial activity. Certain compoundsprovided herein possess potent antibacterial, antiprotozoal, orantifungal activity. Particular compounds provided herein are alsopotent and/or selective inhibitors of prokaryotic DNA synthesis andprokaryotic reproduction. The invention provides anti-microbialcompositions, including pharmaceutical compositions, containing one ormore carrier, diluents, or excipients. The invention providespharmaceutical compositions containing a1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione or related compoundas the only active agent or containing a1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione or related compoundin combination with one or more other active agent, such as one or moreother antimicrobial or antifungal agent. The invention provides methodsfor treating or preventing microbial infections in eukaryotes,preferably animals, by administering an effective amount of a1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione or related compoundto a eukaryote suffering from or susceptible to microbial infection. Theinvention also provides methods of inhibiting microbial growth andsurvival by applying an effective amount of a1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione or relatedcompound.

The invention also provides novel intermediates useful for the for thesynthesis of 1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-diones andrelated compounds. The invention also provides methods of synthesis of1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-diones and relatedcompounds.

BACKGROUND OF THE INVENTION

Antimicrobial compounds are compounds capable of destroying orsuppressing the growth or reproduction of microorganisms, such asbacteria, protozoa, mycoplasma, yeast, and fungi. The mechanisms bywhich antimicrobial compounds act vary. However, they are generallybelieved to function in one or more of the following ways: by inhibitingcell wall synthesis or repair; by altering cell wall permeability; byinhibiting protein synthesis; or by inhibiting synthesis of nucleicacids. For example, beta-lactam antibacterials inhibit the essentialpenicillin binding proteins (PBPs) in bacteria, which are responsiblefor cell wall synthesis. Quinolones act, at least in part, by inhibitingsynthesis of DNA, thus preventing the cell from replicating.

Many attempts to produce improved antimicrobials yield equivocalresults. Indeed, few antimicrobials are produced that are trulyclinically acceptable in terms of their spectrum of antimicrobialactivity, avoidance of microbial resistance, and pharmacology. Thusthere is a continuing need for broad-spectrum antimicrobials, and aparticular need for antimicrobials effective against resistant microbes.

Pathogenic bacteria are known to acquire resistance via several distinctmechanisms including inactivation of the antibiotic by bacterial enzymes(e.g., beta-lactamases that hydrolyze penicillin and cephalosporins);removal of the antibiotic using efflux pumps; modification of the targetof the antibiotic via mutation and genetic recombination (e.g.,penicillin-resistance in Neiserria gonorrhea); and acquisition of areadily transferable gene from an external source to create a resistanttarget (e.g., methicillin-resistance in Staphylococcus aureus). Thereare certain gram-positive pathogens, such as vancomycin-resistantEnterococcus faecium, which are resistant to virtually all commerciallyavailable antibiotics.

Resistant organisms of particular note include methicillin-resistant andvancomycin-resistant Staphylococcus aureus, penicillin-resistantStreptococcus pneumoniae, vancomycin-resistant enterococci,fluoroquinolone-resistant E. coli, cephalosporin-resistant aerobicgram-negative rods and imipenem-resistant Pseudomonas aeruginosa. Theseorganisms are significant causes of nosocomial infections and areclearly associated with increasing morbidity and mortality. Theincreasing numbers of elderly and immunocompromised patients areparticularly at risk for infection with these pathogens. Therefore,there is a large unmet medical need for the development of newantimicrobial agents.

SUMMARY OF THE INVENTION

The invention provides compounds of Formula I and Formula II (shownbelow) and includes 1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dioneand related compounds, which possess antimicrobial activity. Theinvention provides compounds of Formula I and Formula II that possesspotent and/or selective antibacterial, antiprotozoal, or antifungalactivity. The invention also provides anti-bacterial compositionscontaining one or more compounds of Formula I or Formula II, or a salt,solvate, or acylated prodrug of such a compound, and one or morecarriers, excipients, or diluents.

The invention further comprises methods of treating and preventingmicrobial infections, particularly bacterial and protozoal infections byadministering and effective amount of a compound of Formula I or FormulaII to a eukaryote suffering from or susceptible to a microbialinfection. These microbial infections include bacterial infections, forexample E. coli infections, Staphylococcus infections, Salmonellainfections and protozoal infections, for example Chlamydia infections.The invention is particularly includes methods of preventing or treatingmicrobial infections in mammals, including humans, but also encompassesmethods of preventing or treating microbial infections in other animals,including fish, birds, reptiles, and amphibians.

Methods of treatment include administering a compound of Formula I orFormula II alone as the single active agent or administering a compoundof Formula I in combination with one or more other therapeutic agent,such as an antibacterial, an antifungal, an antiviral, an interferon, anefflux-pump inhibitor, a beta-lactamase inhibitor, or another compoundof Formula I or Formula II.

The invention also provides methods of inhibiting microbial growth andsurvival by applying an effective amount of an1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione or relatedcompound. The invention includes, for example, methods of inhibitingmicrobial growth and survival on medical instruments or on surfaces usedfor food preparation by applying a composition containing a compound ofFormula I or Formula II.

Thus, the invention include compounds of Formula I and Formula II

and the pharmaceutically acceptable salts thereof, wherein:

-   -   A₁ is S, O, SO, or SO₂.    -   R₂ is hydrogen, or    -   R₂ is C₁-C₈alkyl, C₂-C₆alkenyl, C₂-C₆alkynyl,        (C₃-C₇cycloalkyl)C₀-C₄carbohydryl,        (C₄-C₇cycloalkenyl)C₀-C₄carbohydryl, (aryl)C₀-C₄carbohydryl, or        (C₂-C₆heterocycloalkyl)C₀-C₄carbohydryl, each of which is        substituted with 0 to 5 substituents independently chosen from        halogen, hydroxy, amino, cyano, nitro, C₁-C₄alkyl, C₁-C₄alkoxy,        C₁-C₂haloalkyl, C₁-C₂haloalkoxy, mono- and di-C₁-C₄alkylamino,        C₂-C₄alkanoyl, C₁-C₄alkylthio, ═NOR₁₀, ═NR₁₀, —O(C═O)R₁₀,        —(C═O)NR₁₀R₁₁, —O(C═O)NR₁₀R₁₁, —(C═O)OR₁₀, —(C═O)NR₁₀OR₁₁,        —NR₁₀(C═O)R₁₁, —NR₁₀(C═O)OR₁₁, —NR₁₀(C═O)OR₁₁R₁₂,        —NR₁₀(C═S)NR₁₁R₁₂, —NR₁₀NR₁₁R₁₂, —SO₃R₁₀, —(S═O)OR₁₀, —SO₂R₁₃,        —SO₂NR₁₀R₁₁, and —NR₁₀SO₂R₁₃; where R₁₀, R₁₁, and R₁₂ are        independently hydrogen, C₁-C₄alkyl, or aryl, and R₁₃ is        C₁-C₄alkyl or aryl.

R₃ is hydrogen, C₁-C₆alkyl, C₁-C₆alkanoyl, mono- ordi-C₁-C₆alkylcarbamate, or C₁-C₆alkylsulfonate; each of which issubstituted with 0 to 3 substituents independently chosen from halogen,hydroxy, amino, cyano, nitro, C₁-C₄alkoxy, mono- and di-C₁-C₄alkylamino,C₁-C₂haloalkyl, and C₁-C₂haloalkoxy.

R₅ is hydrogen, halogen, hydroxy, amino, cyano, nitro, or —NHNH₂, or

R₅ is C₁-C₄alkyl, C₁-C₄alkoxy, mono- or di-(C₁-C₄)alkylamino, mono-, di-or tri-C₁-C₄alkylhydrazinyl, C₂-C₄alkanoyl, C₁-C₄alkylester,C₁-C₂haloalkyl, or C₁-C₂haloalkoxy; each of which is substituted with 0to 3 substituents independently chosen from hydroxy, amino, halogen,oxo, C₁-C₄alkoxy, C₁-C₂haloalkyl, C₁-C₂haloalkoxy, and mono- anddi-C₁-C₄alkylamino.

R₆ is hydrogen, halogen, hydroxy, amino, cyano, C₁-C₄alkyl, C₁-C₄alkoxy,mono- or di-(C₁-C₄)alkylamino, —SO₃R₁₀, —SO₂R₁₀, or —SO₂NR₁₀R₁₁.

R₇is chloro, bromo, iodo, —O(SO₂)CF₃, or —N₂BF₄, or

R₇ is XR_(A) where X is absent, —CH₂—CH₂—, —CH═CH—, —(C═O)—, —(C═O)NH—,or —C≡C—, and R_(A) is C₁-C₆alkyl, C₃-C₇cycloalkyl, C₃-C₇cycloalkenyl,phenyl, a bicyclic 7- to 10-membered saturated, partially unsaturated,or aromatic carbocyclic group, a monocyclic or bicyclic a 5- to10-membered bicyclic saturated, partially unsaturated, or aromaticheterocylic group bound via a carbon atom; each of which R_(A) issubstituted with 0 to 5 substituents independently chosen from (i),(ii), and (iii); where

(i) is chosen from halogen, hydroxy, amino, cyano, and nitro,

(ii) is chosen from C₁-C₆alkyl, C₂-C₆alkenyl, C₂-C₆alkynyl,(C₁-C₆alkoxy)C₀-C₄alkyl, mono- and di-(C₁-C₄)alkylamino, C₁-C₂haloalkyl,C₁-C₂haloalkoxy, (C₃-C₇cycloalkyl)C₀-C₄carbohydryl,(C₃-C₇cycloalkyl)C₀-C₄carbohydryl-O—,(C₄-C₇cycloalkenyl)C₀-C₄carbohydryl, (aryl)C₀-C₆carbohydryl,(aryl)C₁-C₄alkoxy, (C₂-C₆heterocycloalkyl)C₀-C₄carbohydryl,(heteroaryl)C₀-C₆carbohydryl, C₁-C₆alkylthio, ═NOR₁₀, ═NR₁₀,—(C₀-C₄alkyl)(C═O)R₁₀, —(C₀-C₄alkyl)O(C═O)R₁₀,—(C₀-C₄alkyl)(C═O)NR₁₀R₁₁, —(C₀-C₄alkyl)O(C═O)NR₁₀R₁₁,—(C₀-C₄alkyl)(C═O)OR₁₀, —(C₀-C₄alkyl)NR₁₀(C═O)R₁₁,—(C₀-C₄alkyl)NR₁₀(C═O)OR₁₁, —(C₀-C₄alkyl)NR₁₀(C═O)NR₁₁R₁₂,—(C₀-C₄alkyl)NR₁₀(C═O)(C₁-C₄alkyl)NR₁₁(C═O)O—R₁₂,—(C₀-C₄alkyl)NR₁₀(C═S)NR₁₁R₁₂, —(C₀-C₄alkyl)NR₁₀NR₁₁R₁₂,—(C₀-C₄alkyl)N═NR₁₃, —(C₀-C₄alkyl)SO₃R₁₀, —(C₀-C₄alkyl)(S═O)OR₁₀,—(C₀-C₄alkyl)SO₂R₁₃, —(C₀-C₄alkyl)SO₂NR₁₀OR₁₁, and—(C₀-C₄alkyl)NR₁₀SO₂R₁₃; and

(iii) is chosen from —OR_(D), —(C═O)R_(D), —SO₂R_(D), —SO₃R_(D),—NR₁₀SO₂R_(D), where R_(D) is C₁-C₄alkyl, (C₃-C₇cycloalkyl)C₀-C₂alkyl,(C₂-C₆heterocycloalkyl)C₀-C₂alkyl, (aryl)C₀-C₂alkyl, and(heteroaryl)C₀-C₂alkyl;

where each of (ii) and (iii) is substituted with 0 to 3 substituentsindependently chosen from halogen, hydroxy, amino, cyano, nitro, oxo,—COOH, —CONH₂, C₁-C₄alkyl, C₂-C₄alkenyl, C₂-C₄alkynyl, C₁-C₄alkoxy,C₁-C₄alkoxycarbonyl, (C₃-C₇cycloalkyl)C₀-C₄carbohydryl,(C₃-C₇cycloalkyl)C₀-C₄alkoxy, mono- and di-(C₁-C₄)alkylamino,C₁-C₂haloalkyl, C₁-C₂haloalkoxy, C₂-C₄alkanoyl and phenyl.

A₈ is N or CR₈ and R₈ is hydrogen, halogen, hydroxy, amino, cyano,nitro, or —NHNH_(2,) or R₈ is C₁-C₄alkyl, C₁-C₄alkoxy, mono- ordi-(C₁-C₄)alkylamino, mono-, di-, or tri-C₁-C₄ alkylhydrazinyl,C₂-C₄alkanoyl, C₁-C₄alkylester, C₁-C₂haloalkyl, or C₁-C₂haloalkoxy; eachof which is substituted with 0 to 3 substituents independently chosenfrom hydroxy, amino, halogen, oxo, C₁-C₄alkoxy, C₁-C₂haloalkyl,C₁-C₂haloalkoxy, and mono- and di-C₁-C₄alkylamino.

R₉ is C₁-C₈alkyl, C₁-C₄alkoxy, mono- or di-(C₁-C₄)alkylamino,C₂-C₄alkanoyl, C₁-C₂haloalkyl, C₁-C₂haloalkoxy,(C₃-C₇cycloalkyl)C₀-C₄alkyl, or phenyl, each of which is substitutedwith 0 to 3 substituents independently chosen from halogen, hydroxy,amino, cyano, nitro, —COOH, —CONH₂, C₁-C₄alkyl, C₂-C₄alkenyl,C₂-C₄alkynyl, C₁-C₄alkoxy, (C₃-C₇cycloalkyl)C₀-C₄alkyl,(C₃-C₇cycloalkyl)C₀-C₄alkoxy, mono- and di-(C₁-C₄)alkylamino,C₁-C₂haloalkyl, C₁-C₂haloalkoxy, and C₂-C₄alkanoyl.

The invention includes novel intermediates useful for the synthesis ofantimicrobial compounds of Formula I and Formula II. These intermediatesare compounds of Formula I and Formula II in which R₇ is chloro, bromo,iodo, —O(SO₂)CF₃, or —N₂BF₄. The invention provides methods ofsynthesizing compounds of Formula I and Formula II comprising couplingan intermediate of the invention to an appropriate aryl or heteroarylboronic acid, aryl or heteroaryl boronic acid ester, or compoundssubstituted with Li, Mg, B, Al, Si, Zn, Cu, Zr, or Sn at the point ofcoupling.

DETAILED DESCRIPTION OF THE INVENTION

Chemical Description and Terminology

Prior to setting forth the invention in detail, it may be helpful toprovide definitions of certain terms to be used herein. Compounds of thepresent invention are generally described using standard nomenclature.

In certain situations, the compounds of Formula I and Formula II maycontain one or more asymmetric elements such as stereogenic centers,stereogenic axes and the like, e.g. asymmetric carbon atoms, so that thecompounds can exist in different stereoisomeric forms. These compoundscan be, for example, racemates or optically active forms. For compoundswith two or more asymmetric elements, these compounds can additionallybe mixtures of diastereomers. For compounds having asymmetric centers,it should be understood that all of the optical isomers and mixturesthereof are encompassed. In addition, compounds with carbon-carbondouble bonds may occur in Z- and E-forms, with all isomeric forms of thecompounds being included in the present invention. In these situations,the single enantiomers, i.e., optically active forms can be obtained byasymmetric synthesis, synthesis from optically pure precursors, or byresolution of the racemates. Resolution of the racemates can also beaccomplished, for example, by conventional methods such ascrystallization in the presence of a resolving agent, or chromatography,using, for example a chiral HPLC column.

Where a compound exists in various tautomeric forms, the invention isnot limited to any one of the specific tautomers, but rather includesall tautomeric forms.

The present invention is intended to include all isotopes of atomsoccurring in the present compounds. Isotopes include those atoms havingthe same atomic number but different mass numbers. By way of generalexample, and without limitation, isotopes of hydrogen include tritiumand deuterium and isotopes of carbon include ¹¹C, ¹³C, and ¹⁴C.

Certain compounds are described herein using a general formula thatincludes variables, e.g. A₁, R₂, R₃, R₅, R₆, R₇, A₈, and A₉. Unlessotherwise specified, each variable within such a formula is definedindependently of other variables. Thus, if a group is said to besubstituted, e.g. with 0-2 R*, then said group may be substituted withup to two R* groups and R* at each occurrence is selected independentlyfrom the definition of R*. Also, combinations of substituents and/orvariables are permissible only if such combinations result in stablecompounds.

The term “substituted”, as used herein, means that any one or morehydrogens on the designated atom or group is replaced with a selectionfrom the indicated group, provided that the designated atom's normalvalence is not exceeded. When a substituent is oxo (i.e., ═O), imine(e.g. ═NR), or oxime (e.g. ═NOR) then 2 hydrogens on the atom arereplaced. An “oxo,” imine, or oxime substituent on an aromatic group orheteroaromatic group destroys the aromatic character of that group, e.g.a pyridyl substituted with oxo is pyridone. Combinations of substituentsand/or variables are permissible only if such combinations result instable compounds or useful synthetic intermediates. A stable compound orstable structure is meant to imply a compound that is sufficientlyrobust to survive isolation from a reaction mixture, and subsequentformulation into an effective therapeutic agent. Unless otherwisespecified substituents are named into the core structure. For example,it is to be understood that when (cycloalkyl)alkyl is listed as apossible substituent the point of attachment of this substituent to thecore structure is in the alkyl portion.

A dash (“—”) that is not between two letters or symbols is used toindicate a point of attachment for a substituent. For example, —CONH₂ isattached through the carbon atom.

As used herein, “alkyl” is intended to include both branched andstraight-chain saturated aliphatic hydrocarbon groups, having thespecified number of carbon atoms. Thus, the term C₁-C₆ alkyl as usedherein includes alkyl groups having from 1 to about 6 carbon atoms. WhenC₀-C_(n) alkyl is used herein in conjunction with another group, forexample, (aryl)C₀-C₄alkyl, the indicated group, in this case aryl, iseither directly bound by a single covalent bond (C₀), or attached by analkyl chain having the specified number of carbon atoms, in this casefrom 1 to about 4 carbon atoms. Examples of alkyl include, but are notlimited to, methyl, ethyl, n-propyl, isopropyl, n-butyl, t-butyl,n-pentyl, and sec-pentyl. Preferred alkyl groups are lower alkyl groups,those alkyl groups having from 1 to about 8 carbon atoms or 1 to about 6carbon atoms, e.g. C₁-C₈ and C₁-C₆alkyl groups.

“Alkenyl” as used herein, indicates a hydrocarbon chain of either astraight or branched configuration having one or more carbon-carbondouble bond bonds, which may occur at any stable point along the chain.Examples of alkenyl groups include ethenyl and propenyl.

“Alkynyl” as used herein, indicates a hydrocarbon chain of either astraight or branched configuration having one or more triplecarbon-carbon bonds that may occur in any stable point along the chain,such as ethynyl and propynyl.

“Carbohydryl” as used herein, includes both branched and straight-chainhydrocarbon groups, which are saturated or unsaturated, having thespecified number of carbon atoms. When C₀-C_(n)carbohydryl is usedherein in conjunction with another group, for example,(aryl)C₀-C₄carbohydryl, the indicated group, in this case aryl, iseither directly bound by a single covalent bond (C₀), or attached by ancarbohydryl chain, such as an alkyl chain, having the specified numberof carbon atoms, in this case from 1 to about 4 carbon atoms. Examplesinclude C₁-C₆alkyl, such as methyl, or 5-butyl, C₂-C₆alkynyl such andhexynyl, and C₂-C₆ alkenyl, such as 1-propenyl.

“Alkoxy” represents an alkyl group as defined above with the indicatednumber of carbon atoms attached through an oxygen bridge. Examples ofalkoxy include, but are not limited to, methoxy, ethoxy, n-propoxy,i-propoxy, n-butoxy, 2-butoxy, t-butoxy, n-pentoxy, 2-pentoxy,3-pentoxy, isopentoxy, neopentoxy, n-hexoxy, 2-hexoxy, 3-hexoxy, and3-methylpentoxy. In the group “(alkoxy)alkyl” the alkoxy and alkylgroups carry the definitions set forth above and the point of attachmentto the core molecule is in the alkyl.

“Alkanoyl” indicates an alkyl group as defined above, attached through aketo (—(C═O)—) bridge. Alkanoyl groups have the indicated number ofcarbon atoms, with the carbon of the keto group being included in thenumbered carbon atoms. For example a C₂alkanoyl group is an acetyl grouphaving the formula CH₃(C═O)—.

As used herein, the terms “mono- or di-alkylamino” or “mono- anddi-alkylamino” indicate secondary or tertiary alkyl amino groups,wherein the alkyl groups are as defined above and have the indicatednumber of carbon atoms. The point of attachment of the alkylamino groupis on the nitrogen. Examples of mono- and di-alkylamino groups includeethylamino, dimethylamino, and methyl-propyl-amino.

The term “mono- or di-alkylcarbamate” indicates 1 or 2 independentlychosen alkyl groups, as defined above, attached through a carbamate(—O(C═O)NRR) linkage where each R represents an independently chosenalkyl group. Mono-alkylcarbamate groups have the formula (—O(C═O)NHR.

The term “alkylester” indicates and alkyl group as define above attachedthrough an ester linkage in either orientation, i.e. a group of theformula —O(C═O)alkyl or a group of the formula —(C═O)Oalkyl.

The term “mono-, di-, or tri-alkylhydrazinyl” indicates from 1 to 3independently chosen alkyl group as defined above attached through asingle-bonded nitrogen-nitrogen linkage. At least one of the alkylgroups is attached to the terminal nitrogen (the nitrogen not bound tothe core structure). When the term mono-alkylhydrazinyl is used only theterminal nitrogen is alkyl substituted. Examples of alkylhydrazinylgroups include 2-butyl-1-hydrazinyl, 2-butyl-2-methyl-1-hydraazinyl, and1,2-dimethyl-2-propyl-1-hydrazinyl.

The term “alkylthio” indicates an alkyl group as defined above attachedthrough a sulfur linkage, i.e. a group of the formula alkyl-S—. Examplesinclude ethylthio and pentylthio.

As used herein, the term “aryl” indicates aromatic groups containingonly carbon in the aromatic ring or rings. Typical aryl groups contain 1to 3 separate, fused, or pendant rings and from 6 to about 18 ringatoms, without heteroatoms as ring members. When indicated, such arylgroups may be further substituted with carbon or non-carbon atoms orgroups. Such substitution may include fusion to a 5 to 7-memberedsaturated cyclic group that optionally contains 1 or 2 heteroatomsindependently chosen from N, O, and S, to form, for example, a3,4-methylenedioxy-phenyl group. Aryl groups include, for example,phenyl, naphthyl, including 1-naphthyl and 2-naphthyl, and bi-phenyl.

In the term “(aryl)alkyl”, aryl and alkyl are as defined above, and thepoint of attachment is on the alkyl group. This term encompasses, but isnot limited to, benzyl, phenylethyl, and piperonyl. Likewise, in theterm “(aryl)carbohydryl”, aryl and carbohydryl are as defined above andthe point of attachment is on the carbohydryl group, for example aphenylpropen-1-yl group.

The term “carbocyclic group” indicates a 5-6 membered saturated,partially unsaturated, or aromatic ring containing only carbon ringatoms or a 7-10 membered bicyclic saturated, partially unsaturated, oraromatic ring system containing only carbon ring atoms. Unless otherwiseindicated, the carbocyclic ring may be attached to its pendant group atany carbon atom that results in a stable structure. I

“Cycloalkyl” as used herein, indicates a saturated hydrocarbon ring,having the specified number of carbon atoms, usually from 3 to about 8ring carbon atoms, or from 3 to about 7 carbon atoms. Examples ofcycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, orcyclohexyl as well as bridged or caged saturated ring groups such asnorborane or adamantane.

“Cycloalkenyl” as used herein, indicates an unsaturated, but notaromatic, hydrocarbon ring having at least one carbon-carbon doublebond. Cycloalkenyl groups contain from 4 to about 8 carbon atoms,usually from 4 to about 7 carbon atoms. Examples include cyclohexenyland cyclobutenyl.

In the terms “(cycloalkyl)alkyl,” “(cycloalkyl)carbohydryl,” and“(cycloalkyl)alkoxy” the terms cycloalkyl, alkyl, carbohydryl, andalkoxy are as defined above, and the point of attachment is on thealkyl, carbohydryl, or alkoxy group respectively. These terms includeexamples such as cyclopropylmethyl, cyclohexylmethyl,cyclohexylpropenyl, and cyclopentylethyoxy. The term(cycloalkyl)carbohydryl-O— indicates a (cycloalkyl)carbohydryl group asdefined above attached via an oxygen bridge.

In the terms “(cycloalkenyl)alkyl” and “(cycloalkenyl)carbohydryl” theterms cycloalkenyl, alkyl, and carbohydryl are as defined above, and thepoint of attachment is on the alkyl or carbohydryl group respectively.These terms include examples such as cyclobutenylmethyl,cyclohexenylmethyl, and cyclohexylpropenyl.

“Haloalkyl” indicates both branched and straight-chain saturatedaliphatic hydrocarbon groups having the specified number of carbonatoms, substituted with 1 or more halogen atoms, generally up to themaximum allowable number of halogen atoms. Examples of haloalkylinclude, but are not limited to, trifluoromethyl, difluoromethyl,2-fluoroethyl, and penta-fluoroethyl.

“Haloalkoxy” indicates a haloalkyl group as defined above attachedthrough an oxygen bridge.

“Halo” or “halogen” as used herein refers to fluoro, chloro, bromo, oriodo.

As used herein, “heteroaryl” indicates a stable 5- to 7-memberedmonocyclic or 7- to 10-membered bicyclic heterocyclic ring whichcontains at least 1 aromatic ring that contains from 1 to 4, orpreferably from 1 to 3, heteroatoms chosen from N, O, and S, withremaining ring atoms being carbon. When the total number of S and Oatoms in the heteroaryl group exceeds 1, these heteroatoms are notadjacent to one another. It is preferred that the total number of S andO atoms in the heteroaryl group is not more than 2. It is particularlypreferred that the total number of S and O atoms in the aromaticheterocycle is not more than 1. A nitrogen atom in a heteroaryl groupmay optionally be quaternized. When indicated, such heteroaryl groupsmay be further substituted with carbon or non-carbon atoms or groups.Such substitution may include fusion to a 5 to 7-membered saturatedcyclic group that optionally contains 1 or 2 heteroatoms independentlychosen from N, O, and S, to form, for example, a[1,3]dioxolo[4,5-c]pyridyl group. Examples of heteroaryl groups include,but are not limited to, pyridyl, indolyl, pyrimidinyl, pyridizinyl,pyrazinyl, imidazolyl, oxazolyl, furanyl, thiophenyl, thiazolyl,triazolyl, tetrazolyl, isoxazolyl, quinolinyl, pyrrolyl, pyrazolyl,benz[b]thiophenyl, isoquinolinyl, quinazolinyl, quinoxalinyl, thienyl,isoindolyl, and 5,6,7,8-tetrahydroisoquinoline.

In the terms “heteroarylalkyl” and “(heteroaryl)carbohydryl,”heteroaryl, alkyl, and carbohydryl are as defined above, and the pointof attachment is on the alkyl or carbohydryl group respectively. Theseterms include such examples as pyridylmethyl, thiophenylmethyl, and(pyrrolyl)1-ethyl.

The term “heterocycloalkyl” indicates a saturated cyclic groupcontaining from 1 to about 3 heteroatoms chosen from N, O, and S, withremaining ring atoms being carbon. Heterocycloalkyl groups have from 3to about 8 ring atoms, and more typically have from 5 to 7 ring atoms.Examples of heterocycloalkyl groups include morpholinyl, piperazinyl,piperidinyl, and pyrrolidinyl groups. A nitrogen in a heterocycloalkylgroup may optionally be quaternized.

The term “heterocyclic group” indicates a 5-6 membered saturated,partially unsaturated, or aromatic ring containing from 1 to about 4heteroatoms chosen from N, O, and S, with remaining ring atoms beingcarbon or a 7-10 membered bicyclic saturated, partially unsaturated, oraromatic heterocylic ring system containing at least 1 heteroatom in thetwo ring system chosen from N, O, and S and containing up to about 4heteroatoms independently chosen from N, O, and S in each ring of thetwo ring system. Unless otherwise indicated, the heterocyclic ring maybe attached to its pendant group at any heteroatom or carbon atom thatresults in a stable structure. When indicated the heterocyclic ringsdescribed herein may be substituted on carbon or on a nitrogen atom ifthe resulting compound is stable. A nitrogen atom in the heterocycle mayoptionally be quatemized. It is preferred that the total number ofheteroatoms in a heterocyclic groups is not more than 4 and that thetotal number of S and O atoms in a heterocyclic group is not more than2, more preferably not more than 1. Examples of heterocyclic groupsinclude, pyridyl, indolyl, pyrimidinyl, pyridizinyl, pyrazinyl,imidazolyl, oxazolyl, furanyl, thiophenyl, thiazolyl, triazolyl,tetrazolyl, isoxazolyl, quinolinyl, pyrrolyl, pyrazolyl,benz[b]thiophenyl, isoquinolinyl, quinazolinyl, quinoxalinyl, thienyl,isoindolyl, dihydroisoindolyl, 5,6,7,8-tetrahydroisoquinoline,pyridinyl, pyrimidinyl, furanyl, thienyl, pyrrolyl, pyrazolyl,pyrrolidinyl, morpholinyl, piperazinyl, piperidinyl, and pyrrolidinyl.

Additional examples heterocyclic groups include, but are not limited to,phthalazinyl, oxazolyl, indolizinyl, indazolyl, benzothiazolyl,benzimidazolyl, benzofuranyl, benzoisoxolyl, dihydro-benzodioxinyl,oxadiazolyl, thiadiazolyl, triazolyl, tetrazolyl, oxazolopyridinyl,imidazopyridinyl, isothiazolyl, naphthyridinyl, cinnolinyl, carbazolyl,beta-carbolinyl, isochromanyl, chromanonyl, chromanyl,tetrahydroisoquinolinyl, isoindolinyl, isobenzotetrahydrofuranyl,isobenzotetrahydrothienyl, isobenzothienyl, benzoxazolyl,pyridopyridinyl, benzotetrahydrofuranyl, benzotetrahydrothienyl,purinyl, benzodioxolyl, triazinyl, phenoxazinyl, phenothiazinyl, 5pteridinyl, benzothiazolyl, imidazopyridinyl, imidazothiazolyl,dihydrobenzisoxazinyl, benzisoxazinyl, benzoxazinyl,dihydrobenzisothiazinyl, benzopyranyl, benzothiopyranyl, coumarinyl,isocoumarinyl, chromanyl, tetrahydroquinolinyl, dihydroquinolinyl,dihydroquinolinonyl, dihydroisoquinolinonyl, dihydrocoumarinyl,dihydroisocoumarinyl, isoindolinonyl, benzodioxanyl, benzoxazolinonyl,pyrrolyl N-oxide, pyrimidinyl N-oxide, pyridazinyl N-oxide, pyrazinylN-oxide, quinolinyl N-oxide, indolyl N-oxide, indolinyl N oxide,isoquinolyl N-oxide, quinazolinyl N-oxide, quinoxalinyl N-oxide,phthalazinyl N-oxide, imidazolyl N-oxide, isoxazolyl N-oxide, oxazolylN-oxide, thiazolyl N-oxide, indolizinyl N oxide, indazolyl N-oxide,benzothiazolyl N-oxide, benzimidazolyl N-oxide, pyrrolyl N-oxide,oxadiazolyl N-oxide, thiadiazolyl N-oxide, tetrazolyl N-oxide,benzothiopyranyl S-oxide, and benzothiopyranyl S,S-dioxide.

As used herein “Active agents” are compounds that have pharmaceuticalutility, e.g. may be used to treat a patient suffering from a disease orcondition, or may be used prophylacticly to prevent the onset of adisease or condition in a patient, or that may be used to enhance thepharmaceutical activity of other compounds.

“Salts” of the compounds of the present invention include inorganic andorganic acid and base addition salts. The salts of the present compoundscan be synthesized from a parent compound that contains a basic oracidic moiety by conventional chemical methods. Generally, such saltscan be prepared by reacting free acid forms of these compounds with astoichiometric amount of the appropriate base (such as Na, Ca, Mg, or Khydroxide, carbonate, bicarbonate, or the like), or by reacting freebase forms of these compounds with a stoichiometric amount of theappropriate acid. Such reactions are typically carried out in water orin an organic solvent, or in a mixture of the two. Generally,non-aqueous media like ether, ethyl acetate, ethanol, isopropanol, oracetonitrile are preferred, where practicable. Salts of the presentcompounds further include solvates of the compounds and of the compoundsalts.

“Pharmaceutically acceptable salts” includes derivatives of thedisclosed compounds wherein the parent compound is modified by makingnon-toxic acid or base salts thereof, and further refers topharmaceutically acceptable solvates of such compounds and such salts.Examples of pharmaceutically acceptable salts include, but are notlimited to, mineral or organic acid salts of basic residues such asamines; alkali or organic salts of acidic residues such as carboxylicacids; and the like. The pharmaceutically acceptable salts include theconventional non-toxic salts and the quaternary ammonium salts of theparent compound formed, for example, from non-toxic inorganic or organicacids. For example, conventional non-toxic acid salts include thosederived from inorganic acids such as hydrochloric, hydrobromic,sulfuric, sulfamic, phosphoric, nitric and the like; and the saltsprepared from organic acids such as acetic, propionic, succinic,glycolic, stearic, lactic, malic, tartaric, citric, ascorbic, pamoic,maleic, hydroxymaleic, phenylacetic, glutamic, benzoic, salicylic,mesylic, esylic, besylic, sulfanilic, 2-acetoxybenzoic, fumaric,toluenesulfonic, methanesulfonic, ethane disulfonic, oxalic, isethionic,HOOC—(CH₂)n-COOH where n is 0-4, and the like. Lists of additionalsuitable salts may be found, e.g., in Remington's PharmaceuticalSciences, 17th ed., Mack Publishing Company, Easton, Pa., p. 1418(1985).

The term “prodrugs” includes any compounds that become compounds ofFormula I when administered to a mammalian subject, e.g., upon metabolicprocessing of the prodrug. Examples of prodrugs include, but are notlimited to, acetate, formate and benzoate and like derivatives offunctional groups (such as alcohol or amine groups) in the compounds ofFormula I and Formula II.

The term “therapeutically effective amount” of a compound of thisinvention means an amount effective, when administered to a human ornon-human patient, to provide a therapeutic benefit such as anamelioration of symptoms, e.g., an amount effective to decrease thesymptoms of a microbial infection, and preferably an amount sufficientto reduce the symptoms of a bacterial, fungal, or protozoal infection.In certain circumstances a patient having a microbial infection may notpresent symptoms of being infected. Thus a therapeutically effectiveamount of a compound is also an amount sufficient to prevent asignificant increase or significantly reduce the detectable level ofmicroorganism or antibodies against the microorganism in the patient'sblood, serum, other bodily fluids, or tissues. The invention alsoincludes using compounds of Formula I and Formula II in prophylactictherapies. In the context of prophylactic or preventative treatment a“therapeutically effective amount” is an amount sufficient tosignificantly decrease the treated human or non-human patient's risk ofcontracting a microorganism infection. A significant reduction is anydetectable negative change that is statistically significant in astandard parametric test of statistical significance such as Student'sT-test, where p<0.05.

Antimicrobial Compounds

For the purposes of this document, the following numbering system willapply to the core 1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione(when A₁=sulfur) structure or core2H-isoxazolo[5,4-b]quinolizine-3,4-dione (when A₁=oxygen) structure. Thenumbers 1 through 9 refer specifically to positions within the tricyclicring system whereas the letters A, B and C refer to the specific six(rings A and B) or five (ring C) member rings as shown below.

In addition to the compounds of Formula I and Formula II, describedabove the invention also includes compounds of Formula I and Formula IIin which for the variables (e.g. A₁, R₂, R₃, R₄, etc.) one or more ofthe following conditions are met.

The A₁ variable

(a). A₁ is Sulfur; e.g. the invention includes compounds of Formula IIIand Formula IV.

(b). A₁ is SO; e.g. the invention includes compounds of Formula V andFormula VI.

(c). A₁ is SO₂; e.g. the invention includes compounds of Formula VII andFormula VIII.

(d). A₁ is O; e.g. the invention includes compounds and salts of FormulaIX and Formula X.

The R₂ variable:

(a). The invention includes compounds of Formula I in which R₂ ishydrogen, or

-   R₂ is C₁-C₆alkyl or (C₃-C₇cycloalkyl) C₀-C₄alkyl, each of which is    substituted with at least one substituent chosen from hydroxy,    amino, —COOH, —(C═O)NR₁₀OR₁₁, and —CONH₂; and is substituted with 0    to 3 substituents independently chosen from halogen, hydroxy, amino,    cyano, nitro, —COOH, —CONH₂, C₁-C₄alkyl, C₁-C₄alkoxy,    C₁-C₂haloalkyl, C₁-C₂haloalkoxy, and mono- and di-C₁-C₄alkylamino,    and C₂-C₄alkanoyl.

(b). R₂ is hydrogen.

The R₃ variable:

(a). R₃ is C₁-C₆alkyl, C₁-C₆alkanoyl, mono- or di-C₁-C₆alkylcarbamate,or C₁-C₆alkylsulfonate; each of which is substituted with 0 to 3substituents independently chosen from halogen, hydroxy, amino, cyano,nitro, C₁-C₄alkoxy, mono- and di-C₁-C₄alkylamino, C₁-C₂haloalkyl, andC₁-C₂haloalkoxy.

(b). R₃ is C₁-C₆alkyl or C₁-C₆alkanoyl, each of which is substitutedwith 0 to 3 substituents independently chosen from halogen, hydroxy,amino, cyano, C₁-C₂alkoxy, mono- and di-C₁-C₂alkylamino, C₁-C₂haloalkyl,and C₁-C₂haloalkoxy.

(c). R₃ is C₁-C₆alkyl or C₁-C₆alkanoyl.

(d). R₃ is hydrogen.

The R₅ variable:

(a). R₅ is hydrogen, amino, C₁-C₂alkyl, C₁-C₂alkoxy, mono- ordi-(C₁-C₄)alkylamino, or mono- or di-C₁-C₄ alkylhydrazinyl.

(b). R₅ is hydrogen, amino, mono- or di-(C₁-C₂)alkylamino, or mono- ordi-C₁-C₂ alkylhydrazinyl.

(c). R₅ is hydrogen.

The R₆ variable

(a). R₆ is hydrogen, halogen, or amino.

(b). R₆ is fluoro or hydrogen.

The R₇ variable:

(a). R₇ is chloro, bromo, iodo, —O(SO₂)CF₃, or —N₂BF₄.

(b). R₇ is chloro.

(c). R₇ is XR_(A) where X is absent, —CH₂, —CH₂—CH₂—, —CH═CH—, or —C≡C—;and R_(A) is C₁-C₆alkyl, C₃-C₇cycloalkyl, C₃-C₇cycloalkenyl, phenyl, abicyclic 7- to 10-membered saturated, partially unsaturated, or aromaticcarbocyclic group, a monocyclic or bicyclic 5- to 10-membered bicyclicsaturated, partially unsaturated, or aromatic heterocylic group boundvia a carbon atom; each of which R_(A) is substituted with 0 to 5substituents independently chosen from (i), (ii), and (iii).

(d). R₇ is XR_(A) where X is absent, —CH₂, —CH₂—CH₂—, —CH═CH—, or —C≡C—;and R_(A) is a C₁-C₆alkyl, C₃-C₇cycloalkyl, C₃-C₇cycloalkenyl, phenyl,phenyl fused to a 5- or 6-membered heterocycloalkyl ring containing 1 or2 nitrogen or oxygen atoms, indanyl naphthyl, dihydronaphthyl,tetrahydronapthyl, benz[b]thiophenyl, benzofuiranyl, dihydroindolyl,dihydroisoindolyl, dihydroquinolinyl, dihydroisoquinolinyl, furanyl,indolyl, imidazolyl, isoindolyl, isoquinolinyl, isoxazolyl, piperazinyl,piperidinyl, pyrazinyl, pyridyl, pyrimidinyl, pyrrolidinyl, pyrrolyl,pyrazolyl, quinolinyl, quinazolinyl, quinoxalinyl, thienyl, thiazolyl,tetrahydropyridinyl, tetrahydroisoquionolinyl, or tetrahydroquinolinylgroup; each of which is substituted with 0 to 5 substituentsindependently chosen from (i), (ii), and (iii).

(e). R₇ is XR_(A) where X is absent, —CH₂—, —CH₂—CH₂—, —CH═CH—, or—C≡C—; and R_(A) is C₁-C₆alkyl, C₃-C₇cycloalkyl, C₃-C₇cycloalkenyl,phenyl, phenyl fused to a 5- or 6-membered heterocycloalkyl ringcontaining 1 or 2 nitrogen or oxygen atoms, indanyl naphthyl,dihydronaphthyl, tetrahydronapthyl, benz[b]thiophenyl, benzofuranyl,dihydroindolyl, dihydroisoindolyl, dihydroquinolinyl,dihydroisoquinolinyl, furanyl, indolyl, imidazolyl, isoindolyl,isoquinolinyl, isoxazolyl, piperazinyl, piperidinyl, pyrazinyl, pyridyl,pyrimidinyl, pyrrolidinyl, pyrrolyl, pyrazolyl, quinolinyl,quinazolinyl, quinoxalinyl, thienyl, thiazolyl, tetrahydropyridinyl,tetrahydroisoquionolinyl, or tetrahydroquinolinyl; group; each of whichis substituted with 0 to 5 substituents independently chosen from (i),(ii), and (iii), wherein

(i) is chosen from halogen, hydroxy, amino, cyano, and nitro,

(ii) is chosen from C₁-C₄alkyl, C₂-C₄alkenyl, C₁-C₄alkoxy, mono- anddi-(C₁-C₄)alkylamino, C₁-C₂haloalkyl, C₁-C₂haloalkoxy,(C₃-C₇cycloalkyl)C₀-C₂alkyl, (C₃-C₇cycloalkyl)C₀-C₂alkoxy,(phenyl)C₀-C₂alkyl, (phenyl)C₀-C₂alkoxy, (pyrrolidinyl)C₀-C₂alkyl,(piperidinyl)C₀-C₂alkyl, (piperazinyl)C₀-C₂alkyl,(morpholinyl)C₀-C₂alkyl, (thiomorpholinyl)C₀-C₂alkyl, pyridyl,pyrimidinyl, pyrazinyl, furanyl, benzofuranyl, pyrrolyl, pyrazolyl,imidazolyl, thienyl, C₁-C₄alkylthio, ═NOR₁₀, —(C₀-C₄alkyl)(C═O)R₁₀,—(C₀-C₄alkyl)O(C═O)R₁₀, —(C₀-C₄alkyl)(C═O)NR₁₀R₁₁,—(C₀-C₄alkyl)O(C═O)NR₁₀R₁₁, —(C₀-C₄alkyl)(C═O)OR₁₀,—(C₀-C₄alkyl)NR₁₀(C═O)R₁₁, —(C₀-C₄alkyl)NR₁₀(C═O)OR₁₁,—(C₀-C₄alkyl)NR₁₀(C═O)NR₁₁R₁₂,—(C₀-C₄alkyl)NR₁₀(C═O)(C₁-C₄alkyl)NR₁₁(C═O)O—R₁₂,—(C₀-C₄alkyl)NR₁₀(C═S)NR₁₁R₁₂, —(C₀-C₄alkyl)NR₁₀NR₁₁R₁₂,—(C₀-C₄alkyl)N═NR₁₃, —(C₀-C₄alkyl)SO₃R₁₀, —(C₀-C₄alkyl)(S═O)OR₁₀,—(C₀-C₄alkyl)SO2R₁₃, —(C₀-C₄alkyl)SO2NR₁₀R₁₁, and—(C₀-C₄alkyl)NR₁₀SO₂R₁₃; and

(iii) is chosen from —OR_(D), —(C═O)R_(D), —SO₂R_(D), —SO₃R_(D), and—NR₁₀SO₂R_(D), where R_(D) is C₁-C₄alkyl, (C₃-C₇cycloalkyl)C₀-C₂alkyl,(pyrrolidinyl)C₀-C₂alkyl, (piperidinyl)C₀-C₂alkyl,(piperazinyl)C₀-C₂alkyl, (morpholinyl)C₀-C₂alkyl,(thiomorpholinyl)C₀-C₂alkyl, (phenyl)C₀-C₂alkyl, (naphthyl)C₀-C₂alkyl,(pyridyl)C₀-C₂alkyl, (pyrimidinyl)C₀-C₂alkyl, (pyrazinyl)C₀-C₂alkyl,(furanyl)C₀-C₂alkyl, (benz[b]thiophenyl)C₀-C₂alkyl,(benzofuranyl)C₀-C₂alkyl, (quinolinyl)C₀-C₂alkyl,(isoquinolinyl)C₀-C₂alkyl, (quinazolinyl)C₀-C₂alkyl,(isoxazolyl)C₀-C₂alkyl, (indolyl)C₀-C₂alkyl, (dihydroindolyl)C₀-C₂alkyl,(pyrrolyl)C₀-C₂alkyl, (pyrazolyl)C₀-C₂alkyl, (imidazolyl)C₀-C₂alkyl,(thienyl)C₀-C₂alkyl, (isoindolyl)C₀-C₂alkyl, or(dihydroisoindolyl)C₀-C₂alkyl; where each of (ii) and (iii) issubstituted with 0 to 3 substituents independently chosen from halogen,hydroxy, amino, cyano, nitro, oxo, —COOH, —CONH₂, C₁-C₄alkyl,C₂-C₄alkenyl, C₂-C₄alkynyl, C₁-C₄alkoxy, C₁-C₄alkoxycarbonyl,(C₃-C₇cycloalkyl)C₀-C₄alkyl, (C₃-C₇cycloalkyl)C₀-C₄alkoxy, mono- anddi-(C₁-C₄)alkylamino, C₁-C₂haloalkyl, C₁-C₂haloalkoxy, C₂-C₄alkanoyl,and phenyl.

(f) R₇ is XR_(A) where X is absent, —CH₂, —CH₂—CH₂—, —CH═CH—, or —C≡C—;and

R_(A) is a C₁-C₆alkyl, C₃-C₇cycloalkyl, C₃-C₇cycloalkenyl, phenyl,phenyl fused to a 5- or 6-membered heterocycloalkyl ring containing 1 or2 nitrogen or oxygen atoms, indanyl naphthyl, dihydronaphthyl,tetrahydronapthyl, benz[b]thiophenyl, benzofuranyl, dihydroindolyl,dihydroisoindolyl, dihydroquinolinyl, dihydroisoquinolinyl, furanyl,indolyl, imidazolyl, isoindolyl, isoquinolinyl, isoxazolyl, piperazinyl,piperidinyl, pyrazinyl, pyridyl, pyrimidinyl, pyrrolidinyl, pyrrolyl,pyrazolyl, quinolinyl, quinazolinyl, quinoxalinyl, thienyl, thiazolyl,tetrahydropyridinyl, tetrahydroisoquionolinyl, or tetrahydroquinolinyl;group; each of which is substituted with 0 to 5 substituentsindependently chosen from (i), (ii), and (iii), wherein

(i) is chosen from halogen, hydroxy, amino, cyano, and nitro,

(ii) is chosen from C₁-C₄alkyl, C₂-C₄alkenyl, C₁-C₄alkoxy, mono- anddi-(C₁-C₄)alkylamino, C₁-C₂haloalkyl, C₁-C₂haloalkoxy, (C₃-C₇cycloalkyl)C₀-C₂alkyl, (C₃-C₇cycloalkyl)C₀-C₂alkoxy, (phenyl)C₀-C₂alkyl,(phenyl)C₀-C₂alkoxy, pyrrolidinyl, piperidinyl, piperazinyl,morpholinyl, thiomorpholinyl, pyridyl, thienyl, C₁-C₄alkylthio,—(C═O)OR₁₀, and —(C═O)NR₁₀R₁₁; and

(iii) is chosen from —OR_(D), —(C═O)R_(D), —SO₂R_(D), —SO₃R_(D), and—NR₁₀SO₂R_(D), where R_(D) is C₁-C₄alkyl, (C₃-C₇cycloalkyl)C₀-C₂alkyl,piperidinyl, piperazinyl, phenyl, naphthyl, or pyridyl; where each of(ii) and (iii) is substituted with 0 to 3 substituents independentlychosen from halogen, hydroxy, amino, cyano, nitro, oxo, —COOH, —CONH₂,C₁-C₄alkyl, C₁-C₄alkoxy, C₁-C₄alkoxycarbonyl,(C₃-C₇cycloalkyl)C₀-C₂alkyl, mono- and di-(C₁-C₄)alkylamino,C₁-C₂haloalkyl, C₁-C₂haloalkoxy, C₂-C₄alkanoyl, and phenyl.

(g). R₇ is XR_(A) where X is absent, —CH₂, —CH₂—CH₂—, —CH═CH—, or —C≡C—;and

R_(A) is a C₁-C₆alkyl, C₃-C₇cycloalkyl, C₃-C₇cycloalkenyl, phenyl,phenyl fused to a 5- or 6-membered heterocycloalkyl ring containing 1 or2 nitrogen or oxygen atoms, indanyl naphthyl, dihydronaphthyl,tetrahydronapthyl, benz[b]thiophenyl, benzofuranyl, dihydroindolyl,dihydroisoindolyl, dihydroquinolinyl, dihydroisoquinolinyl, furanyl,indolyl, imidazolyl, isoindolyl, isoquinolinyl, isoxazolyl, piperazinyl,piperidinyl, pyrazinyl, pyridyl, pyrimidinyl, pyrrolidinyl, pyrrolyl,pyrazolyl, quinolinyl, quinazolinyl, quinoxalinyl, thienyl, thiazolyl,tetrahydropyridinyl, tetrahydroisoquionolinyl, or tetrahydroquinolinyl;group; each of which is substituted with 0 to 5 substituentsindependently chosen from halogen, hydroxy, amino, cyano, nitro,C₁-C₄alkyl, C₂-C₄alkenyl, C₁-C₄alkoxy, mono- and di-(C₁-C₄)alkylamino,C₁-C₂haloalkyl, C₁-C₂haloalkoxy, (C₃-C₇cycloalkyl)C₀-C₂alkyl,(C₃-C₇cycloalkyl)C₀-C₂alkoxy, (phenyl)C₀-C₂alkyl, (phenyl)C₀-C₂alkoxy,pyrrolidinyl, piperidinyl, piperazinyl, morpholinyl, thiomorpholinyl,pyridyl, thienyl, C₁-C₄alkylthio, —(C═O)OR₁₀, —(C═O)NR₁₀R₁₀; and—OR_(D), —(C═O)R_(D), —SO₂R_(D), —SO₃R_(D), and —NR₁₀SO₂R_(D), whereR_(D) is C₁-C₄alkyl, piperidinyl, phenyl, naphthyl, and pyridyl; andeach R_(D) is substituted with 0 to 3 substituents independently chosenfrom halogen, hydroxy, amino, cyano, C₁-C₂alkyl, C₁-C₂alkoxy, mono- anddi-(C₁-C₂)alkylamino, C₁-C₂haloalkyl, and C₁-C₂haloalkoxy.

(h) R₇ is XR_(A) where X is absent and R_(A) carries the definition setforth in (f) or (g) above.

(i) R₇ is XR_(A) where X is absent, —CH₂, —CH₂—CH₂—, —CH═CH—, or —C≡C—;or in certain embodiments X is absent, and

R_(A) is phenyl or pyridyl, each of which is substituted with 0 to 3substituents independently chosen from halogen, hydroxy, amino, cyano,nitro, C₁-C₄alkyl, C₂-C₄alkenyl, C₁-C₄alkoxy, mono- anddi-(C₁-C₄)alkylamino, C₁-C₂haloalkyl, C₁-C₂haloalkoxy,(C₃-C₇cycloalkyl)C₀-C₂alkyl, (C₃-C₇cycloalkyl)C₀-C₂alkoxy,(phenyl)C₀-C₂alkyl, (phenyl)C₀-C₂alkoxy, pyrrolidinyl, piperidinyl,piperazinyl, morpholinyl, thiomorpholinyl, pyridyl, thienyl,C₁-C₄alkylthio, —(C═O)OR₁₀, and —(C═O)NR₁₀R₁₁; and substituted with 1 or2 substituents independently chosen from —OR_(D), —(C═O)R_(D),—SO₂R_(D), —SO₃R_(D), and —NR₁₀SO₂R_(D), where R_(D) is C₁-C₄alkyl,piperidinyl, phenyl, naphthyl, or pyridyl; and each R_(D) is substitutedwith 0 to 3 substituents independently chosen from halogen, hydroxy,amino, cyano, C₁-C₂alkyl, C₁-C₂alkoxy, mono- and di-(C₁-C₂)alkylamino,C₁-C₂haloalkyl, and C₁-C₂haloalkoxy.

(j) R₇ is XR_(A) where X is absent and R_(A) is a phenyl, pyridyl,pyrimidinyl, furanyl, isoquinolinyl, quinolinyl, indolyl, pyrrolyl,isoindolyl, dihydroisoindolyl, tetrahydroisoquinolinyl,tetrahydroisoquinolinyl, or thienyl group, each of which is substitutedwith 0 to 3 substituents independently chosen from halogen, hydroxy,amino, cyano, C₁-C₄alkyl, C₁-C₄alkoxy, mono- and di-(C₁-C₄)alkylamino,C₁-C₂haloalkyl, and C₁-C₂haloalkoxy.

(k) R₇ is XR_(A) where X is absent and R_(A) is phenyl, pyrid-3-yl,pyrid-4-yl, pyrimidin-5-yl, furan-3-yl, quinolin-3-yl, quinolin-5-yl,quinolin-6-yl, isoindol-5-yl, tetrahydroisoquinolin-5-yl,tetrahydroisoquinolin-6-yl, tetrahydroisoquinolin-7-yl,tetrahydroisoquinolin-8-yl, or indol-5-yl, each of which is substitutedwith 0 to 2 substituents independently chosen from halogen, hydroxy,amino, C₁-C₂alkyl, and C₁-C₂alkoxy.

(l) R₇ is XR_(A) where X is absent and R_(A) is phenyl, pyridyl-3-yl orpyrid-4-yl, each of which is substituted with 1 or 2 substituentsindependently chosen from fluoro, amino, hydroxy, cyano, and methyl.

(m) R₇ is XR_(A) where X is absent and R_(A) istetrahydroisoquinolin-5-yl, tetrahydroisoquinolin-6-yl,tetrahydroisoquinolin-7-yl, or tetrahydroisoquinolin-8-yl, each of whichis substituted with 0 to 3 substituents independently chosen fromC₁-C₃alkyl.

(n) R₇ is XR_(A) where X is absent and R_(A) istetrahydroisoquinolin-6-yl optionally substituted at each of the 1, 2,and 3 positions with methyl.

(o) R₇ is XR_(A) where X is absent and R_(A) is isoindol-5-ylsubstituted with 0 to 3 independently chosen C₁-C₃alkyl substituents.

(p) R₇ is XR_(A) where X is absent and R_(A) is isoindol-5-yl optionallysubstituted at each of the 1, 2, or 3 positions with methyl.

(q) R₇ is XR_(A) where X is absent, —CH₂, —CH₂—CH₂—, —CH═CH—, or —C≡C—;or in certain embodiments X is absent, and R_(A) is phenyl fused to a 5-or 6-membered heterocycloalkyl ring containing 1 or 2 nitrogen or oxygenatoms, where R_(A) is substituted with 0 to 3 substituents independentlychosen from (i), (ii), and (iii).

(r) R₇ is XR_(A) where X is absent, —CH₂, —CH₂—CH₂—, —CH═CH—, or —C≡C—;or in certain embodiments X is absent, R_(A) is phenyl fused to a 5- or6-membered heterocycloalkyl ring containing 1 or 2 nitrogen or oxygenatoms, where R_(D) is substituted with 0 to 2 substituents independentlychosen from halogen, hydroxy, amino, C₁-C₂alkyl, and C₁-C₂alkoxy.

The A₈ variable

(a). A₈ is nitrogen.

(b). A₈ is CR₈.

(c). A₈ is CR₈ and R₈ is hydrogen, halogen, C₁-C₂alkyl, C₁-C₂alkoxy,trifluoromethyl, or trifluoromethoxy.

(d). A₈ is CR₈ and R₈ is hydrogen or methyl.

The R₉ variable:

(a). R₉ is C₁-C₈alkyl, C₁-C₄alkoxy, mono- or di-(C₁-C₄)alkylamino,C₂-C₄alkanoyl, C₁-C₂haloalkyl, C₁-C₂haloalkoxy,(C₃-C₇cycloalkyl)C₀-C₄alkyl, or phenyl, each of which is substitutedwith 0 to 3 substituents independently chosen from halogen, hydroxy,amino, C₁-C₂alkyl, C₁-C₂alkoxy, mono- and di-(C₁-C₂)alkylamino,C₁-C₂haloalkyl, and C₁-C₂haloalkoxy.

(b). R₉ is C₁-C₄alkyl, cyclopropyl, or phenyl, each of which issubstituted with 0 to 3 substituents independently chosen from halogen,hydroxy, amino, C₁-C₂alkyl, C₁-C₂alkoxy, mono- and di-(C₁-C₂)alkylamino,C₁-C₂haloalkyl, and C₁-C₂haloalkoxy.

(c). R₉ is C₁-C₄alkyl or cyclopropyl, or R₉ is phenyl substituted with 2substituents chosen from halogen, hydroxy, amino, C₁-C₂alkyl,C₁-C₂alkoxy, mono- and di-(C₁-C₂)alkylamino, C₁-C₂haloalkyl, andC₁-C₂haloalkoxy.

(d). R₉ is ethyl, t-butyl, cyclopropyl, or 2,4-difluorophenyl.

(e). R₉ is cyclopropyl.

The invention includes compounds of Formula I and Formula II in whichthe variables A₁, R₂, R₃, R₅, R₆, R₇, A₈, and R₉ carry any combinationof the definitions set forth for these variables above that results in astable compound.

Certain compounds of Formula I and Formula II exhibit potentantibacterial, antifungal, and/or antiprotozoal activity. Particularcompounds of the invention exhibit Minimum Inhibitory Concentrations(MIC) of 64 μg/ml or less against Staphyloccocus aureus and/orEschericia coli in a standard assay for determining the MIC of acompound against these bacteria, such as the assay provided in Example 9below. Preferred compounds of Formula I and II exhibit MIC values of 10μg/ml or less against Staphyloccocus aureus and/or Eschericia coli. Morepreferred compound of the Formula I and II exhibit MIC values of 4 μg/mlor less, or even more preferably 1 μg/ml or less, against Staphyloccocusaureus and/or Eschericia coli.

Certain compounds of Formula I and Formula II are selectiveantimicrobial agents; having the ability to kill or inhibit the growthor reproduction of pathogenic microbial organisms, while having littleor no effect on the cells of fish, amphibians, reptiles, birds, ormammals. The selectivity of compounds of Formula I and Formula II may beassessed by determining the CC₅₀ (the concentration at which 50% of thecells are killed) for cultured cells of a higher animal, such as a fish,reptiles, amphibian, bird, or mammal. Certain compounds of the inventionexhibit a CC₅₀ of greater that 100 micromolar for mammalian cells.Certain compounds of the invention exhibit a CC50 of greater than 100micromolar for cultured human hepatocytes, and also exhibit MIC valuesof 64 μg/ml or less, preferably 10 μg/ml or less, or more preferably 4μg/ml or less, or still more preferably 1 μg/ml or less againstStaphyloccocus aureus and/or Eschericia coli.

Without wishing to be bound to any particular theory it is believed thatthe antimicrobial properties of compounds of Formula I and Formula IIare due to the ability to these compounds to inhibit the activity ofmicrobial DNA gyrases while having little or no effect on the analogousenzyme, Topoisomerase II, present in higher organisms. Certain preferredcompounds of the invention are 100-fold or more selective for bacterialDNA gyrases than for mammalian, particularly human, Topoisomerase II.

Synthetic Intermediates

The invention includes novel intermediates useful for the synthesis ofantimicrobial compounds of Formula I and Formula II. Coupling reactionsoccur between R′-M and R″-Y in the presence of catalyst Q, where M isLi, Mg, B, Al, Si, Zn, Cu, Zr, or Sn; where Y is I, Br, Cl, —O(SO₂)CF₃,or —N₂BF₄; Q is Fe, Ni, Cu, Pd, or Rh; and R′ and R″ are organicmolecules to which M and Y are bound. M may also be Boron, disubstitutedwith OH, OG, or G, where G is an optionally substituted straight,branched, or cyclic alkyl group, or other suitable group; Y is Br, andwhere Q is Pd. A general review of this chemistry can be found in Tamao,K and Miyaura, N. Topics in Current Chemistry 219: 1-9 (2002). A reviewof the use of coupling reagents in which M is Boron with a listing ofpotential boronates, palladium catalysts, and reaction conditions can befound in Miyaura, N. Topics in Current Chemistry 219: 11-59 (2002).

Thus the invention includes intermediates of Formula XI and Formula XII:

Wherein A₁, A₈, R₂, R₃, R₅, R₆, and R₉ carry the definitions set forthabove and Y is I, Br, Cl, —O(SO₂)CF₃, or —N₂BF₄. Here compounds ofFormula XI and Formula XII play the role of R″—Y intermediate in thecoupling reaction between R′-M and R″—Y. These intermediates are coupledto compounds of the Formula R′-M where R′ is a group R.

R is XR_(A) where X is absent, —CH₂—CH₂—, —CH═CH—, —(C═O)—, —(C═ONH)—,or —C≡C—, and R_(A) is C₁-C₆alkyl, C₃-C₇cycloalkyl, C₃-C₇cycloalkenyl,phenyl, a bicyclic 7- to 10-membered saturated, partially unsaturated,or aromatic carbocyclic group, a monocyclic or bicyclic a 5- to10-membered bicyclic saturated, partially unsaturated, or aromaticheterocylic group bound via a carbon atom; each of which R_(A) issubstituted with 0 to 5 substituents independently chosen from (i),(ii), and (iii); where

(i) is chosen from halogen, hydroxy, amino, cyano, and nitro,

(ii) is chosen from C₁-C₆alkyl, C₂-C₆alkenyl, C₂-C₆alkynyl,(C₁-C₆alkoxy)C₀-C₄alkyl, mono- and di-(C₁-C₄)alkylamino, C₁-C₂haloalkyl,C₁-C₂haloalkoxy, (C₃-C₇cycloalkyl)C₀-C₄carbohydryl,(C₃-C₇cycloalkyl)C₀-C₄carbohydryl-O—,(C₄-C₇cycloalkenyl)C₀-C₄carbohydryl, (aryl)C₀-C₆carbohydryl,(aryl)C₁-C₄alkoxy, (C₂-C₆heterocycloalkyl)C₀-C₄carbohydryl,(heteroaryl)C₀-C₆carbohydryl, C₁-C₆alkylthio, ═NOR₁₀, ═NR₁₀,—(C₀-C₄alkyl)(C═O)R₁₀, —(C₀-C₄alkyl)O(C═O)R₁₀,—(C₀-C₄alkyl)(C═O)NR₁₀R₁₁, —(C₀-C₄alkyl)O(C═O)NR₁₀R₁₀,—(C₀-C₄alkyl)(C═O)OR₁₀, —(C₀-C₄alkyl)NR₁₀(C═O)R₁₁,—(C₀-C₄alkyl)NR₁₀(C═O)OR₁₁, —(C₀-C₄alkyl)NR₁₀(C═O)NR₁₁R,—(C₀-C₄alkyl)NR₁₀(C═O)(C₁-C₄alkyl)NR₁₁(C═O)O—R₁₂,—(C₀-C₄alkyl)NR₁₀(C═S)NR₁₁R₁₂, —(C₀-C₄alkyl)NR₁₀NR₁₁R₁₂,—(C₀-C₄alkyl)N═NR₁₃, —(C₀-C₄alkyl)SO₃R₁₀, —(C₀-C₄alkyl)(S═O)OR₁₀,—(C₀-C₄alkyl)SO₂R₁₃, —(C₀-C₄alkyl)SO2NR₁₀R₁₁, and—(C₀-C₄alkyl)NR₁₀SO₂R₁₃; and

-   -   (iii) is chosen from —OR_(D), —(C═O)RD, —SO₂RD, —SO₃RD,        —NR₁₀SO2RD, where RD is C1-C4alkyl, (C₃-C₇cycloalkyl)C₀-C₂alkyl,        (C2-C6heterocycloalkyl)C₀-C₂alkyl, (aryl)C₀-C₂alkyl, and        (heteroaryl)C₀-C₂alkyl;

where each of (ii) and (iii) is substituted with 0 to 3 substituentsindependently chosen from halogen, hydroxy, amino, cyano, nitro, oxo,—COOH, —CONH2, C1-C4alkyl, C2-C4alkenyl, C2-C4alkynyl, C1-C4alkoxy,C1-C4alkoxycarbonyl, (C₃-C₇cycloalkyl)C₀-C₄carbohydryl,(C₃-C₇cycloalkyl)C₀-C₄alkoxy, mono- and di-(C1-C4)alkylamino,C₁-C₂haloalkyl, C₁-C₂haloalkoxy, C2-C4alkanoyl and phenyl.

M is Li, Mg, B, Al, Si, Zn, Cu, Zr, or Sn; or M is Boron, disubstitutedwith OH, OG, or G, where G is an optionally substituted straight,branched, or cyclic alkyl group, an optionally substituted aryl orarylalkyl group, or other suitable group.

The invention also includes intermediates of Formula XIII and FormulaXIV.

In Formula XIII and XIV A₁, A₈, R₂, R₃, R₅, R₆, and R₉ carry thedefinitions set forth above and M is Li, Mg, B, Al, Si, Zn, Cu, Zr, orSn; or M is Boron, disubstituted with OH, OG, or G, where G is anoptionally substituted straight, branched, or cyclic alkyl group, anoptionally substituted aryl or arylalkyl group, or other suitable group.Here compounds of Formula XIII and Formula XIV play the role of R′-M inthe coupling reaction. These intermediates are coupled to compounds ofthe Formula R″-Y where Y is I, Br, Cl, —O(SO₂)CF₃, or —N₂BF₄ and R″carries the definition set forth above for R.

Pharmaceutical Preparations

Compounds and salts of Formula I and Formula II can be administered asthe neat chemical, but are preferably administered as a pharmaceuticalcomposition or formulation. Accordingly, the invention providespharmaceutical formulations comprising a compound or pharmaceuticallyacceptable salt of Formula I or Formula II, together with one or morepharmaceutically acceptable carrier, excipients, adjuvant, diluent, orother ingredient.

Compounds of general Formula I and Formula II may be administeredorally, topically, parenterally, by inhalation or spray, sublingually,transdermally, via buccal administration, rectally, as an ophthalmicsolution, or by other means, in dosage unit formulations containingconventional non-toxic pharmaceutically acceptable carriers, excipients,adjuvants, and vehicles.

In addition to the subject compound, the compositions of the inventionmay contain a pharmaceutically acceptable carrier, one or morecompatible solid or liquid filler, diluents or encapsulating substances,which are suitable for administration to an animal. Carriers must be ofsufficiently high purity and sufficiently low toxicity to render themsuitable for administration to the animal being treated. The carrier canbe inert or it can possess pharmaceutical benefits of its own. Theamount of carrier employed in conjunction with the compound issufficient to provide a practical quantity of material foradministration per unit dose of the compound.

Exemplary pharmaceutically acceptable carriers or components thereof aresugars, such as lactose, glucose and sucrose; starches, such as cornstarch and potato starch; cellulose and its derivatives, such as sodiumcarboxymethyl cellulose, ethyl cellulose, and methyl cellulose; powderedtragacanth; malt; gelatin; talc; solid lubricants, such as stearic acidand magnesium stearate; calcium sulfate; vegetable oils, such as peanutoil, cottonseed oil, sesame oil, olive oil, and corn oil; polyols suchas propylene glycol, glycerine, sorbitol, mannitol, and polyethyleneglycol; alginic acid; emulsifiers, such as the TWEENS; wetting agents,such sodium lauryl sulfate; coloring agents; flavoring agents; tabletingagents, stabilizers; antioxidants; preservatives; pyrogen-free water;isotonic saline; and phosphate buffer solutions.

In particular, pharmaceutically acceptable carriers for systemicadministration include sugars, starches, cellulose and its derivatives,malt, gelatin, talc, calcium sulfate, vegetable oils, synthetic oils,polyols, alginic acid, phosphate buffer solutions, emulsifiers, isotonicsaline, and pyrogen-free water. Preferred carriers for parenteraladministration include propylene glycol, ethyl oleate, pyrrolidone,ethanol, and sesame oil.

Optional active agents may be, included in a pharmaceutical composition,which do not substantially interfere with the activity of the compoundof the present invention.

Effective concentrations of one or more of the compounds of theinvention including pharmaceutically acceptable salts, esters or otherderivatives thereof are mixed with a suitable pharmaceutical carrier,excipients, adjuvant, or vehicle. In instances in which the compoundsexhibit insufficient solubility, methods for solubilizing compounds maybe used. Such methods are known to those of skill in this art, andinclude, but are not limited to, using cosolvents, such asdimethylsulfoxide (DMSO), using surfactants, such as Tween, ordissolution in aqueous sodium bicarbonate. Derivatives of the compounds,such as salts of the compounds or prodrugs of the compounds may also beused in formulating effective pharmaceutical compositions.

Upon mixing or addition of the compound(s) of Formula I and/or FormulaII, the resulting mixture may be a solution, suspension, emulsion or thelike. The form of the resulting mixture depends upon a number offactors, including the intended mode of administration and thesolubility of the compound in the chosen carrier or vehicle. Theeffective concentration sufficient for ameliorating the symptoms of thedisease, disorder or condition treated and may be empiricallydetermined.

The pharmaceutical compositions containing compounds of general FormulaI and/or Formula II may be in a form suitable for oral use, for example,as tablets, troches, lozenges, aqueous or oily suspensions, dispersiblepowders or granules, emulsions, hard or soft capsules, or syrups orelixirs. Compositions intended for oral use may be prepared according toany method known to the art for the manufacture of pharmaceuticalcompositions and such compositions may contain one or more agents, suchas sweetening agents, flavoring agents, coloring agents and preservingagents, in order to provide pharmaceutically elegant and palatablepreparations.

Oral formulations contain between 0.1 and 99% of a compound of theinvention and usually at least about 5% (weight %) of a compound of thepresent invention. Some embodiments contain from about 25% to about 50%or from 5% to 75% of a compound of invention.

Liquids Formulations

Compounds of the invention can be incorporated into oral liquidpreparations such as aqueous or oily suspensions, solutions, emulsions,syrups, or elixirs, for example. Moreover, formulations containing thesecompounds can be presented as a dry product for constitution with wateror other suitable vehicle before use. Such liquid preparations cancontain conventional additives, such as suspending agents (e.g.,sorbitol syrup, methyl cellulose, glucose/sugar, syrup, gelatin,hydroxyethyl cellulose, carboxymethyl cellulose, aluminum stearate gel,and hydrogenated edible fats), emulsifying agents (e.g., lecithin,sorbitan monsoleate, or acacia), non-aqueous vehicles, which can includeedible oils (e.g., almond oil, fractionated coconut oil, silyl esters,propylene glycol and ethyl alcohol), and preservatives (e.g., methyl orpropyl p-hydroxybenzoate and sorbic acid).

Orally administered compositions also include liquid solutions,emulsions, suspensions, powders, granules, elixirs, tinctures, syrups,and the like. The pharmaceutically acceptable carriers suitable forpreparation of such compositions are well known in the art. Oralformulations may contain preservatives, flavoring agents, sweeteningagents, such as sucrose or saccharin, taste-masking agents, and coloringagents.

Typical components of carriers for syrups, elixirs, emulsions andsuspensions include ethanol, glycerol, propylene glycol, polyethyleneglycol, liquid sucrose, sorbitol and water. Syrups and elixirs may beformulated with sweetening agents, for example glycerol, propyleneglycol, sorbitol or sucrose. Such formulations may also contain ademulcent.

Suspensions

For a suspension, typical suspending agents include methylcellulose,sodium carboxymethyl cellulose, Avicel RC-591, tragacanth and sodiumalginate; typical wetting agents include lecithin and polysorbate 80;and typical preservatives include methyl paraben and sodium benzoate.

Aqueous suspensions contain the active material(s) in admixture withexcipients suitable for the manufacture of aqueous suspensions. Suchexcipients are suspending agents, for example sodiumcarboxymethylcellulose, methylcellulose, hydropropylmethylcellulose,sodium alginate, polyvinylpyrrolidone, gum tragacanth and gum acacia;dispersing or wetting agents; may be a naturally-occurring phosphatide,for example, lecithin, or condensation products of an alkylene oxidewith fatty acids, for example polyoxyethylene stearate, or condensationproducts of ethylene oxide with long chain aliphatic alcohols, forexample heptadecaethyleneoxycetanol, or condensation products ofethylene oxide with partial esters derived from fatty acids and ahexitol such as polyoxyethylene sorbitol substitute, or condensationproducts of ethylene oxide with partial esters derived from fatty acidsand hexitol anhydrides, for example polyethylene sorbitan substitute.The aqueous suspensions may also contain one or more preservatives, forexample ethyl, or n- propyl p-hydroxybenzoate.

Oily suspensions may be formulated by suspending the active ingredientsin a vegetable oil, for example peanut oil, olive oil, sesame oil orcoconut oil, or in a mineral oil such as liquid paraffin. The oilysuspensions may contain a thickening agent, for example beeswax, hardparaffin or cetyl alcohol. Sweetening agents such as those set forthabove, and flavoring agents may be added to provide palatable oralpreparations. These compositions may be preserved by the addition of ananti-oxidant such as ascorbic acid.

Emulsions

Pharmaceutical compositions of the invention may also be in the form ofoil-in-water emulsions. The oily phase may be a vegetable oil, forexample olive oil or peanut oil, or a mineral oil, for example liquidparaffin or mixtures of these. Suitable emulsifying agents may benaturally-occurring gums, for example gum acacia or gum tragacanth,naturally-occurring phosphatides, for example soy bean, lecithin, andesters or partial esters derived from fatty acids and hexitol,anhydrides, for example sorbitan monoleate, and condensation products ofthe said partial esters with ethylene oxide, for example polyoxyethylenesorbitan monoleate.

Dispersible Powders

Dispersible powders and granules suitable for preparation of an aqueoussuspension by the addition of water provide the active ingredient inadmixture with a dispersing or wetting agent, suspending agent and oneor more preservatives. Suitable dispersing or wetting agents andsuspending agents are exemplified by those already mentioned above.

Tablets and Capsules

Tablets typically comprise conventional pharmaceutically compatibleadjuvants as inert diluents, such as calcium carbonate, sodiumcarbonate, mannitol, lactose and cellulose; binders such as starch,gelatin and sucrose; disintegrants such as starch, alginic acid andcroscarmelose; lubricants such as magnesium stearate, stearic acid andtalc. Glidants such as silicon dioxide can be used to improve flowcharacteristics of the powder mixture. Coloring agents, such as the FD&Cdyes, can be added for appearance. Sweeteners and flavoring agents, suchas aspartame, saccharin, menthol, peppermint, and fruit flavors, areuseful adjuvants for chewable tablets. Capsules (including time releaseand sustained release formulations) typically comprise one or more soliddiluents disclosed above. The selection of carrier components oftendepends on secondary considerations like taste, cost, and shelfstability.

Such compositions may also be coated by conventional methods, typicallywith pH or time-dependent coatings, such that the subject compound isreleased in the gastrointestinal tract in the vicinity of the desiredtopical application, or at various times to extend the desired action.Such dosage forms typically include, but are not limited to, one or moreof cellulose acetate phthalate, polyvinylacetate phthalate,hydroxypropyl methylcellulose phthalate, ethyl cellulose, Eudragitcoatings, waxes and shellac.

Formulations for oral use may also be presented as hard gelatin capsuleswherein the active ingredient is mixed with an inert solid diluent, forexample, calcium carbonate, calcium phosphate or kaolin, or as softgelatin capsules wherein the active ingredient is mixed with water or anoil medium, for example peanut oil, liquid paraffin or olive oil.

Injectable and Parenteral Formulations

Pharmaceutical compositions may be in the form of a sterile injectableaqueous or oleaginous suspension. This suspension may be formulatedaccording to the known art using those suitable dispersing or wettingagents and suspending agents that have been mentioned above. The sterileinjectable preparation may also be sterile injectable solution orsuspension in a non-toxic parentally acceptable diluent or solvent, forexample as a solution in 1,3-butanediol. Among the acceptable vehiclesand solvents that may be employed are water, Ringer's solution, andisotonic sodium chloride solution. In addition, sterile, fixed oils areconventionally employed as a solvent or suspending medium. For thispurpose any bland fixed oil may be employed including synthetic mono- ordiglycerides. In addition, fatty acids such as oleic acid are useful inthe preparation of injectables.

Compounds of Formula I and Formula II may be administered parenterallyin a sterile medium. Parenteral administration includes subcutaneousinjections, intravenous, intramuscular, intrathecal injection orinfusion techniques. The drug, depending on the vehicle andconcentration used, can either be suspended or dissolved in the vehicle.Advantageously, adjuvants such as local anesthetics, preservatives andbuffering agents can be dissolved in the vehicle. In compositions forparenteral administration the carrier comprises at least about 90% byweight of the total composition.

Suppositories

Compounds of Formula I and Formula II may also be administered in theform of suppositories for rectal administration of the drug. Thesecompositions can be prepared by mixing the drug with a suitablenon-irritating excipient that is solid at ordinary temperatures butliquid at the rectal temperature and will therefore melt in the rectumto release the drug. Such materials are cocoa butter and polyethyleneglycols.

Topical Formulations

Compounds of the invention may be formulated for local or topicalapplication, such as for topical application to the skin and mucousmembranes, such as in the eye, in the form of gels, creams, and lotionsand for application to the eye or for intracistemal or intraspinalapplication. Topical compositions of the present invention may be in anyform including, for example, solutions, creams, ointments, gels,lotions, milks, cleansers, moisturizers, sprays, skin, patches, and thelike.

Such solutions may be formulated as 0.01%-10% isotonic solutions, pHabout 5-7, with appropriate salts. Compounds of the invention may alsobe formulated for transdermal administration as a transdermal patch.

Topical compositions containing the active compound can be admixed witha variety of carrier materials well known in the art, such as, forexample, water, alcohols, aloe vera gel, allantoin, glycerine, vitamin Aand E oils, mineral oil, propylene glycol, PPG-2 myristyl propionate,and the like.

Other materials suitable for use in topical carriers include, forexample, emollients, solvents, humectants, thickeners and powders.Examples of each of these types of materials, which can be used singlyor as mixtures of one or more materials, are as follows:

Emollients, such as stearyl alcohol, glyceryl monoricinoleate, glycerylmonostearate, propane-1,2-diol, butane-1,3-diol, mink oil, cetylalcohol, iso-propyl isostearate, stearic acid, iso-butyl palmitate,isocetyl stearate, oleyl alcohol, isopropyl laurate, hexyl laurate,decyl oleate, octadecan-2-ol, isocetyl alcohol, cetyl palmitate,dimethylpolysiloxane, di-n-butyl sebacate, iso-propyl myristate,iso-propyl palmitate, iso-propyl stearate, butyl stearate, polyethyleneglycol, triethylene glycol, lanolin, sesame oil, coconut oil, arachisoil, castor oil, acetylated lanolin alcohols, petroleum, mineral oil,butyl myristate, isostearic acid, palmitic acid, isopropyl linoleate,lauryl lactate, myristyl lactate, decyl oleate, and myristyl myristate;propellants, such as propane, butane, iso-butane, dimethyl ether, carbondioxide, and nitrous oxide; solvents, such as ethyl alcohol, methylenechloride, iso-propanol, castor oil, ethylene glycol monoethyl ether,diethylene glycol monobutyl ether, diethylene glycol monoethyl ether,dimethyl sulphoxide, dimethyl formamide, tetrahydrofuran; humectants,such as glycerin, sorbitol, sodium 2-pyrrolidone-5-carboxylate, solublecollagen, dibutyl phthalate, and gelatin; and powders, such as chalk,talc, fullers earth, kaolin, starch, gums, colloidal silicon dioxide,sodium polyacrylate, tetra alkyl ammonium smectites, trialkyl arylammonium smectites, chemically modified magnesium aluminium silicate,organically modified montmorillonite clay, hydrated aluminium silicate,fumed silica, carboxyvinyl polymer, sodium carboxymethyl cellulose, andethylene glycol monostearate.

The compounds of the invention may also be topically administered in theform of liposome delivery systems, such as small unilamellar vesicles,large unilamellar vesicles, and multilamellar vesicles. Liposomes can beformed from a variety of phospholipids, such as cholesterol,stearylamine or phosphatidylcholines.

Other Formulations

Other compositions useful for attaining systemic delivery of the subjectcompounds include sublingual, buccal and nasal dosage forms. Suchcompositions typically comprise one or more of soluble filler substancessuch as sucrose, sorbitol and mannitol, and binders such as acacia,microcrystalline cellulose, carboxymethyl cellulose and hydroxypropylmethylcellulose. Glidants, lubricants, sweeteners, colorants,antioxidants and flavoring agents disclosed above may also be included.

Compositions for inhalation typically can be provided in the form of asolution, suspension or emulsion that can be administered as a drypowder or in the form of an aerosol using a conventional propellant(e.g., dichlorodifluoromethane or trichlorofluoromethane).

Additional Components

The compositions of the present invention may also optionally comprisean activity enhancer. The activity enhancer can be chosen from a widevariety of molecules that function in different ways to enhanceantimicrobial effects of compounds of the present invention. Particularclasses of activity enhancers include skin penetration enhancers andabsorbtion enhancers.

Pharmaceutical compositions of the invention may also contain additionalactive agents can be chosen from a wide variety of molecules, which canfunction in different ways to enhance the antimicrobial or therapeuticeffects of a compound of the present invention. These optional otheractive agents, when present, are typically employed in the compositionsof the invention at a level ranging from about 0.01% to about 15%. Someembodiments contain from about 0.1% to about 10% by weight of thecomposition. Other embodiments contain from about 0.5% to about 5% byweight of the composition.

Packaged Formulations

The invention includes packaged pharmaceutical formulations. Suchpackaged formulations include a pharmaceutical composition containingone or more compounds or salts of Formula I in a container andinstructions for using the composition to treat an animal (typically ahuman patient) suffering from a microorganism infection) or prevent amicroorganism infection in an animal.

In all of the foregoing the compounds of the invention can beadministered alone or as mixtures, and the compositions may furtherinclude additional drugs or excipients as appropriate for theindication.

Methods of Treatment

The invention includes methods of preventing and treating microorganisminfections, particularly bacterial and protozoal infections, byadministering a therapeutically effective amount of one or morecompounds of Formula I and of Formula II to an animal at risk for amicroorganism infection or having a microorganism infection. The animalmay be a fish, amphibian, reptile or bird, but is preferably a mammal.Methods of treating and preventing microorganism infections in livestockanimals, companion animals, and human patients are particularlypreferred.

The compounds disclosed herein are useful for preventing and treatingbacterial infections in animals. Furthermore compounds of the inventionmay be used to treat a variety of conditions not attributed to bacterialinfections. These include diseases and disorders caused fungalinfections, mycoplasma infections, protozoal infections, or otherconditions involving infectious organisms.

In some circumstances an effective amount of a compound of Formula I orFormula II may be an amount sufficient to reduce the symptoms of themicroorganism infection. Alternatively an effective amount of a Compoundof Formula I may be an amount sufficient to significantly reduce theamount of microorganism or antibodies against the detectable in apatient's tissues or bodily fluids.

Methods of treatment also include inhibiting microorganism replicationin vivo, in an animal at risk for a microorganism infection or sufferingfrom such an infection, by administering a sufficient concentration of acompound of Formula I or Formula II to inhibit bacterial survival invitro. By “sufficient concentration” of a compound administered to thepatient is meant the concentration of the compound available in theanimal's system to prevent or combat the infection. Such a concentrationby be ascertained experimentally, for example by assaying bloodconcentration of the compound, or theoretically, by calculatingbioavailability. The amount of a compound sufficient to inhibitbacterial survival in vitro may be determined with a conventional assayfor bacterial survival such as the Minimum Inhibitory Concentration(MIC) Assay disclosed in Example 9, which follows.

The invention also includes using compounds of Formula I and Formula IIin prophylactic therapies. In the context of prophylactic orpreventative treatment an effective amount of a compound of theinvention is an amount sufficient to significantly decrease the treatedanimal's risk of contracting a microorganism infection.

Compounds of the invention are particularly useful for treating andpreventing infectious disorders. These include for example: ocularinfections such as conjunctivitis; urinary tract and genital infections,such as complicated urinary tract infections, acute urinary tract andgenital infections, such as pyelonephritis, cervical gonococcalinfections, cystitis, urethral chlamydial infections, cervicalchlamydial infections, urethral gonococcal infections, and prostatitis,respiratory infections, such as lower respiratory tract infections,acute sinusitis, acute exacerbations of chronic bronchitis,community-acquired pneumonia, and nosocomial pneumonia, skin infections,such as skin-structure infections, impetigo, folliculitis, boils,scalded skin syndrome, and cellulites, and other infections such as boneinfections, joint infections, infectious diarrhea, typhoid fever,intra-abdominal infections, gynecologic infections, including toxicshock syndrome, pelvic infections, and post-surgical infections.

The disclosed compounds are useful for treating infections caused by thefollowing microorganisms:

Aerobic Gram-positive Microorganisms: Including but not limited toEnterococcus faecalis, Enterococcus faecium, Staphylococcus aureus,Staphylococcus epidermidis, Staphylococcus saprophyticus, Streptococcuspneumoniae, Streptococcus pyogenes, Staphylococcus haemolyticus, andStaphylococcus hominis.

Aerobic Gram-negative Microorganisms: Including but not limited toCampylobacter jejuni, Citrobacter diversus, Citrobacter freundii,Enterobacter cloacae, Escherichia coli, Haemophilus influenzae,Haemophilus parainfluenzae, Klebsiella pneumoniae, Moraxellacatarrhalis, Morganella morganii, Neisseria gonorrhoeae, Proteusmirabilis, Proteus vulgaris, Providencia rettgeri, Providencia stuartii,Pseudomonas aeruginosa, Stenotrophomonas maltophila, Salmonella typhi,Serratia marcescens, Shigella boydii, Shigella dysenteriae, Shigellaflexneri, Shigella sonnei. Acinetobacter Iwoffi, Aeromonas hydrophila,Edwardsiella tarda, Enterobacter aerogenes, Klebsiella oxytoca,Legionella pneumophila, Pasteurella multocida, Salmonella enteritidis,Vibrio cholerae, Vibrio parahaemolyticus, Vibrio vulnificus, Yersiniaenterocolitica and H. Pylorii.

Non-bacterial microorganisms: Mycoplasma, Legionella and Chlamydia.

Dosage levels of the order of from about 0.1 mg to about 140 mg perkilogram of body weight per day are useful in the treatment of theabove-indicated conditions (about 0.5 mg to about 7 g per patient perday). The amount of active ingredient that may be combined with thecarrier materials to produce a single dosage form will vary dependingupon the host treated and the particular mode of administration. Dosageunit forms will generally contain between from about 1 mg to about 500mg of an active ingredient.

Frequency of dosage may also vary depending on the compound used and theparticular disease treated. However, for treatment of most infectiousdisorders, a dosage regimen of 4 times daily or less is preferred and adosage regimen of 1 or 2 times daily is particularly preferred.

It will be understood, however, that the specific dose level for anyparticular patient will depend upon a variety of factors including theactivity of the specific compound employed, the age, body weight,general health, sex, diet, time of administration, route ofadministration, and rate of excretion, drug combination and the severityof the particular disease undergoing therapy.

Combination Administration

The compounds of the invention may also be useful in combination withother pharmaceutically active agents such as antibacterial agents,antiviral agents, antifungal agents, anti-inflammatories, interferon,efflux-pump inhibitors, and beta-lactamase inhibitors. Antibiotic agentsinclude any molecule that tends to prevent, inhibit or destroy life andas such, includes anti-bacterial agents, anti-fungicides, anti-viralagents, and anti-parasitic agents.

A composition comprising a compound or salt of Formula I or Formula IIin combination with one or more other antibacterial agent, antiprotozoalagent, antifungal agent, antiviral agent, interferon, efflux-pumpinhibitor, or beta-lactamase inhibitor is provided herein.

Pharmaceutical compositions of the invention include single dosage formscontaining of a compound of Formula I and/or Formula II and one or moreother active agent, dosage forms containing more than one compound ofFormula I and/or Formula II, and separate. administration of a compoundof Formula I and/or Formula II with another active agent.

The following active agents, which are useful in combinations of theinvention, may be isolated from an organism that produces the agent orsynthesized by methods known to those of ordinary skill in the art ofmedicinal chemistry or purchased from a commercial source.

Anti-bacterial antibiotic agents include, but are not limited to,penicillins, cephalosporins, carbacephems, cephamycins, carbapenems,monobactams, aminoglycosides, glycopeptides, quinolones, tetracyclines,macrolides, and fluoroquinolones (see Table below). Examples ofantibiotic agents include, but are not limited to, Penicillin G (CASRegistry No.: 61-33-6); Methicillin (CAS Registry No.: 61-32-5);Nafcillin (CAS Registry No.: 147-52-4); Oxacillin (CAS Registry No.:66-79-5); Cloxacillin (CAS Registry No.: 61-72-3); Dicloxacillin (CASRegistry No.: 3116-76-5); Ampicillin (CAS Registry No.: 69-53-4);Amoxicillin (CAS Registry No.: 26787-78-0); Ticarcillin (CAS RegistryNo.: 34787-01-4); Carbenicillin (CAS Registry No.: 4697-36-3);Mezlocillin (CAS Registry No.: 51481-65-3); Azlocillin (CAS RegistryNo.: 37091-66-0); Piperacillin (CAS Registry No.: 61477-96-1); Imipenem(CAS Registry No.: 74431-23-5); Aztreonam (CAS Registry No.:78110-38-0); Cephalothin (CAS Registry No.: 153-61-7); Cefazolin (CASRegistry No.: 25953-19-9); Cefaclor (CAS Registry No.: 70356-03-5);Cefamandole formate sodium (CAS Registry No.: 42540-40-9); Cefoxitin(CAS Registry No.: 35607-66-0); Cefuroxime (CAS Registry No.:55268-75-2); Cefonicid (CAS Registry No.: 61270-58-4); Cefmetazole (CASRegistry No.: 56796-20-4); Cefotetan (CAS Registry No.: 69712-56-7);Cefprozil (CAS Registry No.: 92665-29-7); Loracarbef (CAS Registry No.:121961-22-6); Cefetamet (CAS Registry No.: 65052-63-3); Cefoperazone(CAS Registry No.: 62893-19-0); Cefotaxime (CAS Registry No.:63527-52-6); Ceftizoxime (CAS Registry No.: 68401-81-0); Ceftriaxone(CAS Registry No.: 73384-59-5); Ceftazidime (CAS Registry No.:72558-82-8); Cefepime (CAS Registry No.: 88040-23-7); Cefixime (CASRegistry No.: 79350-37-1); Cefpodoxime (CAS Registry No.: 80210-62-4);Cefsulodin (CAS Registry No.: 62587-73-9); Fleroxacin (CAS Registry No.:79660-72-3); Nalidixic acid (CAS Registry No.: 389-08-2); Norfloxacin(CAS Registry No.: 70458-96-7); Ciprofloxacin (CAS Registry No.:85721-33-1); Ofloxacin (CAS Registry No.: 82419-36-1); Enoxacin (CASRegistry No.: 74011-58-8); Lomefloxacin (CAS Registry No.: 98079-51-7);Cinoxacin (CAS Registry No.: 28657-80-9); Doxycycline (CAS Registry No.:564-25-0); Minocycline (CAS Registry No.: 10118-90-8); Tetracycline (CASRegistry No.: 60-54-8); Amikacin (CAS Registry No.: 37517-28-5);Gentamicin (CAS Registry No.: 1403-66-3); Kanamycin (CAS Registry No.:8063-07-8); Netilmicin (CAS Registry No.: 56391-56-1); Tobramycin (CASRegistry No.: 32986-56-4); Streptomycin (CAS Registry No.: 57-92-1);Azithromycin (CAS Registry No.: 83905-01-5); Clarithromycin (CASRegistry No.: 81103-11-9); Erythromycin (CAS Registry No.: 114-07-8);Erythromycin estolate (CAS Registry No.: 3521-62-8); Erythromycin ethylsuccinate (CAS Registry No.: 41342-53-4); Erythromycin glucoheptonate(CAS Registry No.: 23067-13-2); Erythromycin lactobionate (CAS RegistryNo.: 3847-29-8); Erythromycin stearate (CAS Registry No.: 643-22-1);Vancomycin (CAS Registry No.: 1404-90-6); Teicoplanin (CAS Registry No.:61036-64-4); Chloramphenicol (CAS Registry No.: 56-75-7); Clindamycin(CAS Registry No.: 18323-44-9); Trimethoprim (CAS Registry No.:738-70-5); Sulfamethoxazole (CAS Registry No.: 723-46-6); Nitrofurantoin(CAS Registry No.: 67-20-9); Rifampin (CAS Registry No.: 13292-46-1);Mupirocin (CAS Registry No.: 12650-69-0); Metronidazole (CAS RegistryNo.: 443-48-1); Cephalexin (CAS Registry No.: 15686-71-2); Roxithromycin(CAS Registry No.: 80214-83-1); Co-amoxiclavuanate; combinations ofPiperacillin and Tazobactam; and their various salts, acids, bases, andother derivatives.

Anti-fingal agents include but are not limited to Amphotericin B,Candicidin, Dermostatin, Filipin, Fungichromin, Hachimycin, Hamycin,Lucensomycin, Mepartricin, Natamycin, Nystatin, Pecilocin, Perimycin,Azaserine, Griseofulvin, Oligomycins, Neomycin, Pyrrolnitrin, Siccanin,Tubercidin, Viridin, Butenafine, Naftifine, Terbinafine, Bifonazole,Butoconazole, Chlordantoin, Chlormidazole, Cloconazole, Clotrimazole,Econazole, Enilconazole, Fenticonazole, Flutrimazole, Isoconazole,Ketoconazole, Lanoconazole, Miconazole, Omoconazole, Oxiconazole,Sertaconazole, Sulconazole, Tioconazole, Tolciclate, Tolindate,Tolnaftate, Fluconawle, Itraconazole, Saperconazole, Terconazole,Acrisorcin, Amorolfine, Biphenamine, Bromosalicylchloranilide,Buclosamide, Calcium Propionate, Chlorphenesin, Ciclopirox, Cloxyquin,Coparaffinate, Diamthazole, Exalamide, Flucytosine, Halethazole,Hexetidine, Loflucarban, Nifuratel, Potassium Iodide, Propionic Acid,Pyrithione, Salicylanilide, Sodium Propionate, Sulbentine,Tenonitrozole, Triacetin, Ujothion, Undecylenic Acid, and ZincPropionate.

Antiviral agents include, but are not limited to, Acyclovir, Cidofovir,Cytarabine, Dideoxyadenosine, Didanosine, Edoxudine, Famciclovir,Floxuridine, Ganciclovir, Idoxuridine, Inosine Pranobex, Lamivudine,MADU, Penciclovir, Sorivudine, Stavudine, Trifluridine, Valacyclovir,Vidarabine, ZaIcitabine, Zidovudine, Acemannan, Acetylleucine,Amantadine, Amidinomycin, Delavirdine, Foscamet, Indinavir,Interferon-alpha, Interferon-beta, Interferon-gamma, Kethoxal, Lysozyme,Methisazone, Moroxydine, Nevirapine, Podophyllotoxin, Ribavirin,Rimantadine, Ritonavir2, Saquinavir, Stailimycin, Statolon,Tromantadine, and Xenazoic Acid.

Antiinflammatory agents include, but are not limited to, Enfenamic Acid,Etofenamate, Flufenamic Acid, Isonixin, Meclofenamic Acid, MefenamicAcid, Niflumic Acid, Talniflumate, Terofenamate, Tolfenamic Acid,Aceclofenac, Acemetacin, Alclofenac, Amfenac, Amtolmetin Guacil,Bromfenac, Bufexamac, Cinmetacin, Clopirac, Diclofenac, Etodolac,Felbinac, Fenclozic Acid, Fentiazac, Glucametacin, Ibufenac,Indomethacin, Isofezolac, Isoxepac, Lonazolac, Metiazinic Acid,Mofezolac, Oxametacine, Pirazolac, Proglumetacin, Sulindac, Tiaramide,Tolmetin, Tropesin, Zomepirac, Bumadizon, Butibufen, Fenbufen, Xenbucin,Clidanac, Ketorolac, Tinoridine, Alminoprofen, Benoxaprofen,Bermoprofen, Bucloxic Acid, Carprofen, Fenoprofen, Flunoxaprofen,Flurbiprofen, Ibuprofen, Ibuproxam, Indoprofen, Ketoprofen, Loxoprofen,Naproxen, Oxaprozin, Piketoprofen, Pirprofen, Pranoprofen, ProtizinicAcid, Suprofen, Tiaprofenic Acid, Ximoprofen, Zaltoprofen, Difenamizole,Epirizole, Apazone, Benzpiperylon, Feprazone, Mofebutazone, Morazone,Oxyphenbutazone, Phenylbutazone, Pipebuzone, Propyphenazone,Ramifenazone, Suxibuzone, Thiazolinobutazone, Acetaminosalol, Aspirin,Benorylate, Bromosaligenin, Calcium Acetylsalicylate, Diflunisal,Etersalate, Fendosal, Gentisic Acid, Glycol Salicylate, ImidazoleSalicylate, Lysine Acetylsalicylate, Mesalamine, Morpholine Salicylate,I-Naphthyl Salicylate, Olsalazine, Parsalmide, Phenyl Acetylsalicylate,Phenyl Salicylate, Salacetamide, Salicylamide O-Acetic Acid,Salicylsulfuric Acid, Salsalate, Sulfasalazine, Ampiroxicam, Droxicam,Isoxicam, Lomoxicam, Piroxicam, Tenoxicam, epsilon-AcetamidocaproicAcid, S-Adenosylmethionine, 3-Amino-4-hydroxybutyric Acid, Amixetrine,Bendazac, Benzydamine, alpha-Bisabolol, Bucolome, Difenpiramide,Ditazol, Emorfazone, Fepradinol, Guaiazulene, Nabumetone, Nimesulide,Oxaceprol, Paranyline, Perisoxal, Proquazone, Superoxide Dismutase,Tenidap, Zileuton, 21-Acetoxypregnenolone, Alclometasone, Algestone,Amcinonide, Beclomethasone, Betamethasone, Budesonide, Chloroprednisone,Clobetasol, Clobetasone, Clocortolone, Cloprednol, Corticosterone,Cortisone, Cortivazol, Deflazacort, Desonide, Desoximetasone,Dexamethasone, Diflorasone, Diflucortolone, Difluprednate, Enoxolone,Fluazacort, Flucloronide, Flumethasone, Flunisolide, FluocinoloneAcetonide, Fluocinonide, Fluocortin Butyl, Fluocortolone,Fluorometholone, Fluperolone Acetate, Fluprednidene Acetate,Fluprednisolone, Flurandrenolide, Fluticasone Propionate, Formocortal,Halcinonide, Halobetasol Propionate, Halometasone, Halopredone Acetale,Hydrocortamate, Hydrocortisone, Loteprednol Etabonale, Mazipredone,Medrysone, Meprednisone, Methylprednisolone, Mometasone Furoate,Paramethasone, Prednicarbate, Prednisolone, Prednisolone25-Diethylamino-acetate, Prednisolone Sodium Phosphate, Prednisone,Prednival, Prednylidene, Rimexolone, Tixocortol, Triamcinolone,Triamcinolone Acetonide, Triamcinolone Benetonide, and TriamcinoloneHexacetonide.

Compounds of the invention may be combined with one or more Betalactamase inhibitor when used in combination with a beta-lactam classantibiotic, such as penicillin or cephalosporins. Beta-lactamaseinhibitors include, but are not limited to Clavulanic acid, Sulbactam,Sultamacillin, and Tazobactam.

Compounds of the invention may also be combined with one or more effluxpump inhibitor, such as a quinazolinone efflux pump inhibitors,d-ornithine-d-homophenylalanine-3-aminoquinoline,Phe-Arg-b-naphthylamide, propafenone, a phenothiazine or thioxantheneefflux pump inhibitor, 1-aza-9-oxafluorenes,N-[4-[2-(3,4-dihydro-6,7-dimethoxy-2(1H)-isoquinolinyl)ethyl]phenyl]-9,10-dihydro-5-methoxy-9-oxo-4-Acridinecarboxamide,reserpine, Milbemycin, Cinchonine, Verapamil,L-phenylalanyl-N-2-naphthalenyl-L-Argininamide (and analogs),5′-methoxyhydnocarpin-D, methylxanthines, FK506, a cyclosporine effluxpump inhibitor, Nocardamine and other siderophores, Amiodarone,Cyclosporin A, Ro11-2933 (DMDP), Quinidine, and the optical isomers ofPropranolol, Quinine (SQ1) and Quinidine, Quinine-10,11-epoxide,Quercetin, Amitriptyline, Taxuspine C derivatives, Emodin, MC-002434;Agosterol A; Pheophorbide; pyridoquinolines such as2,2′-[(2,8,10-trimethylpyrido[3,2-g]quinoline-4,6-diyl)bis(oxy)]bis[N,N-dimethyl-ethanamine,Gitonavir, and Gemfibrozil.

Synthesis of Compounds

The starting materials used in preparing the compounds of the inventionare known, made by known methods, or are commercially available.

It is recognized that the skilled artisan in the art of organicchemistry can readily carry out standard manipulations of organiccompounds without further direction. Examples of such manipulations arediscussed in standard texts such as J. March, Advanced OrganicChemistry, John Wiley & Sons, 1992. Those skilled in the art of organicsynthesis will recognize that reactions conditions and startingmaterials will vary on occasion from those set forth in the syntheticexamples included herein depending of the desired final product.

The skilled artisan will readily appreciate that certain reactions arebest carried out when other functionalities are masked or protected inthe compound, thus increasing the yield of the reaction and/or avoidingany undesirable side reactions. Often, the skilled artisan utilizesprotecting groups to accomplish such increased yields or to avoid theundesired reactions. These reactions are found in the literature and arealso well within the scope of the skilled artisan. Examples of many suchmanipulations can be found in, for example, T. Greene, Protecting Groupsin Organic Synthesis, John Wiley & Sons, 1981.

The compounds of the invention may have one or more chiral center. As aresult, one may selectively prepare one optical isomer, includingdiastereomers and enantiomers, over another, for example by chiralstarting materials, catalysts or solvents, or may prepare bothstereoisomers or both optical isomers, including diastereomers andenantiomers at once (a racemic mixture). Since the compounds of theinvention may exist as racemic mixtures, mixtures of optical isomers,including diastereomers and enantiomers, or stereoisomers may beseparated using known methods, such as through the use of, for example,chiral salts and chiral chromatography.

In addition, it is recognized that one optical isomer, including adiastereomer and enantiomer, or a stereoisomer, may have favorableproperties over the other. When a racemic mixture is discussed herein,it is clearly contemplated to include both optical isomers, includingdiastereomers and enantiomers, or one stereoisomer substantially free ofthe other.

The invention also includes also includes all energetically accessibleconformational and torsional isomers of the compounds disclosed.

When the substituent R₇ in a compound of Formula I or Formula II isattached via an unsaturated aliphatic group, for example when R₇ is(phenyl)C₂C6alkenyl, all geometric isomers of the compound are included.This invention is further illustrated by the following examples thatshould not be construed as limiting. The contents of all references,patents and published patent applications cited throughout thisapplication are incorporated herein by reference.

EXAMPLES

Abbreviations

The following abbreviations are used in the reaction schemes andsynthetic examples, which follow. This list in not meant to be anall-inclusive list of abbreviations used in the application asadditional standard abbreviations, which are readily understood by thoseskilled in the art of organic synthesis, may also be used in thesynthetic schemes and examples:

-   -   DMF—N,N-Dimethylformamide    -   DMSO—Dimethylsulfoxide    -   LDA—Lithium diisopropylamide    -   Pd(PPh₃)₄-Tetrakis(triphenylphosphine)palladium(0)    -   TFA Trifluoroacetic acid        General Methods

All nonaqueous reactions are performed under an atmosphere of dry argongas (99.99%) using oven- or flame-dried glassware. All isolated productsare stored for prolonged periods under an atmosphere of argon at −20° C.Microwave-assisted syntheses are conducted in a commercial microwavereactor (Discover System, CEM Corporation). The progress of reactions ismonitored using thin-layer chromatography on glass plates coated withMerck silica gel 60 (F254). Flash column chromatography is performed onMerck silica gel 60 (230-400 mesh). Melting points are recorded on anElectrothermal Model IA9100 digital melting point apparatus. NMR spectraare recorded at ambient temperature using a Bruker Avance 300spectrometer (¹H at 300.1 MHz, ¹³C at 75.5 MHz, and ¹⁵N at 30.4 MHz).The chemical shifts for ¹H and ¹³C are reported in parts per million (δ)relative to external tetramethylsilane and are referenced to signals ofresidual protons in the deuterated solvent. The chemical shifts for ¹⁵Nare reported in parts per million (δ) relative to external liquidammonia. Assignment of 1H and ¹³C NMR data is based on extensivetwo-dimensional correlation experiments (¹H—¹H COSY, ¹H—¹³C HMQC, ¹H—¹³CHMBC, and ¹H—¹H NOESY spectra recorded using standard pulse sequences)and the usual principles of NMR spectroscopy (the magnitudes of couplingconstants and chemical shifts). Analytical HPLC is performed using a YMCPack Pro C18 50×4.6 mm 5 μm column with an isocratic elution of 0.24 minat 95:5 H₂O:CH₃CN containing 0.1% TFA followed by a 20-min lineargradient elution from 95:5 to 5:95 at a flow rate of 1.0 mL/min with UVdetection at 254 nm. Low-resolution mass spectra are recorded on aThermo Finnigan Surveyor MSQ instrument (operating in API mode) equippedwith a Gilson liquid chromatograph. High-resolution mass spectrometricanalyses (ESI using sodium iodide as internal standard) are performed atthe W. M. Keck Foundation Biotechnology Resource Laboratory (YaleUniversity, New Haven, Conn.). Elemental analyses are performed atAtlantic Microlab, Inc. (Norcross, Ga.) and Prevalere Life Sciences,Inc. (Whitesboro, N.Y.).

Example 1 Preparation of10-ethyl-8-phenyl-2H-isothiazolo[5,4-b]quinolizine-3,4-dione (7)

Step 1. Diethyl 2-[bis(4-methoxybenzylsulfanyl)methylene]malonate (1)

A solution of dimethylformamide (50 mL) containing diethyl malonate(12.78 g, 0.08 mol) is added slowly to an ice-cooled suspension ofsodium hydride (60% dispersion in mineral oil, 6.53 g, 0.16 mol) indimethylformamide (500 mL). The rate of addition of diethyl malonate issuch as to maintain the temperature of the reaction mixture below 5° C.After the addition of diethyl malonate is complete, the gray reactionmixture continues to stir at 0° C. for 15 min. Carbon disulfide (14.5mL, 0.24 mol) is then added dropwise to give a red mixture that isallowed to warm to room temperature with stirring for 1 h. A solution ofdimethylformamide (25 mL) containing 4-methoxybenzyl chloride (25.00 g,0.16 mol) is added dropwise to the reaction mixture at room temperature.The resulting orange mixture is allowed to warm to room temperature withstirring for 15 h, diluted with water (500 mL), and extracted with ethylacetate (4×500 mL). The combined extracts are concentrated under reducedpressure (˜1 L), washed with water (3×150 mL), washed with brine (3×150mL), dried over magnesium sulfate, and evaporated under reduced pressureto give crude 1. The crude product is purified by flash columnchromatography (eluting with ethyl acetate/hexanes=1:4; Rf 0.21) to give1 as a yellow solid. mp 57-59° C. ¹H NMR (CDCl3): δ 1.27 (t, J=7.0 Hz,3H, CO₂CH₂CH₃), 3.79 (s, 3H, OCH₃), 4.07 (s, 2H, SCH₂), 4.23 (q, J=7.0Hz, 2H, CO₂CH₂CH₃), 6.83 (m, 2H, H-3/H-5), 7.20 (m, 2H, H-2/H-6).¹³C{¹H} NMR (CDCl₃): δ 14.0 (CO₂CH₂CH₃), 40.3 (SCH₂), 55.3 (OCH₃), 61.4(CO₂CH₂CH₃), 114.1 (aromatic C-3/C-5), 128.1 (═C—CO₂CH₂CH₃ or C—CH₂S—),128.7 (═C—CO₂CH₂CH₃ or C—CH₂S—), 130.4 (aromatic C-2/C-6), 155.6(—CH₂S—C═), 159.1 (aromatic C-4, C—OCH₃), 163.9 (CO₂CH₂CH₃). LCMS m/zcalcd for C₂₄H₂₈O₆S₂ 476 ([M+]). found 477 ([M+H]+). HRMS m/z calcd forC₂₄H₂₈NaO₆S₂ 499.1225 ([M+Na]+). found 499.1230. Anal. Calcd forC₂₄H₂₈O₆S₂: C, 60.48; H, 5.92; S, 13.46. Found: C, 60.51; H, 5.97; S,13.19.

Step 2. 4-Bromo-2-propylpyridine (2)

Prepared by a method analogous to the preparation of5-Chloro-2-propylpyridine, described previously (Li, Q., et al. J. Med.Chem. 39: 3070-3088 (1996)). The crude product is purified by flashcolumn chromatography (eluting with ethyl acetate/hexanes=1:2; Rf 0.56)to give 2 as an amber liquid.

¹H NMR (CDCl₃): δ 0.97 (t, J=7.5 Hz, 3H, CH₂CH₂CH₃), 1.74 (m, 2H,CH₂CH₂CH₃), 2.75 (m, 2H, CH₂CH₂CH₃), 7.11 (dd, J=5.5 Hz, 1.5 Hz, 1H,pyridyl H-5), 7.16 (d, J=1.5 Hz, 1H, pyridyl H-3), 8.42 (d, J=5.5 Hz,1H, pyridyl H-6). ¹³C {¹H} NMR (CDCl₃): δ 13.8 (CH₂CH₂CH₃), 22.8(CH₂CH₂CH₃), 40.1 (CH₂CH₂CH₃), 121.3 (CH, C-5), 123.0 (CH, C-3), 144.1(C-4, C-C1), 150.1 (CH, C-6), 164.0 (C-2). LCMS m/z calcd for C₈H₁₀ClN155 ([M+]). found 156 ([M+H]+). HRMS m/z calcd for C8H₁₁ClN 156.0580([M+H]+). found 156.0578.

Step 3. (E)-diethyl2-(1-(4-methoxybenzylthio)-2-(4-bromopyridin-2-yl)but-1-enyl)malonate(3)

Lithium diisopropylamide (LDA) is formed by dropwise addition ofn-butyllithium (1.6 M in hexanes, 10.1 mL, 16.2 mmol) to a stirredsolution of diisopropylamine (2.3 mL, 16.4 mmol) in tetrahydrofuran(25.0 mL) at −78° C. The resulting solution is stirred at −78° C. for 5min, 0° C. for 15 min, and then recooled to −78° C. A solution of4-bromo-2-propylpyridine (2, 2.94 g, 14.7 mmol) in tetrahydrofuran (25.0mL) is added dropwise to this solution over a period of 30 min. Theresulting deep red suspension is stirred at −78° C. for 1 h, after whichtime a solution of diethyl2-[bis(4-methoxybenzylsulfanyl)methylene]malonate (1, 7.03 g, 14.8 mmol)in tetrahydrofuran (40.0 mL) is added dropwise over a period of 15 min.The resulting yellow solution is stirred at −78° C. for 1.5 h, −15° C.for 1.5 h, and room temperature for 30 min to give a dark orangesolution. The reaction mixture is quenched with a saturated aqueoussolution of ammonium chloride (100 mL) and extracted with diethyl ether(2×100 mL). The combined organic extracts are dried over magnesiumsulfate and evaporated under reduced pressure to give 3 an orange oil.The product is purified by flash column chromatography (eluting with 5%ethyl acetate in methylene chloride.

¹H NMR (DMSO-d6): δ 0.76 (t, J=7.5 Hz, 3H, CH₂CH₃), 1.17 (t, J=7.0 Hz,6H, 2×CO₂CH₂CH₃), 2.81 (q, J=7.5 Hz, 2H, CH₂CH₃), 3.74 (s, 3H, OCH₃),4.01 (s, 2H, SCH₂), 4.10 (q, J=7.0 Hz, 4H, 2×CO₂CH₂CH₃), 4.41 (s, 1H,CH(CO₂CH₂CH₃)₂), 6.90 (m, 2H, benzylic H-3/H-5), 7.28 (m, 2H, benzylicH-2/H-6), 7.44 (d, J=2.0 Hz, 1H, pyridyl H-3), 7.52 (dd, J=5.5 Hz, 2.0Hz, 1H, pyridyl H-5), 8.56 (d, J=5.5 Hz, 1H, pyridyl H-6). ¹³C{¹H} NMR(DMSO-d6): δ 12.4 (CH₂CH₃), 13.9 (CH(CO₂CH₂CH₃)₂), 28.3 (CH₂CH₃), 39.5(SCH₂), 55.0 (OCH₃), 58.5 (CH(CO₂CH₂CH₃)₂), 61.1 (CH(CO₂CH₂CH₃)₂), 113.9(benzylic C-3/C-5), 122.9 (pyridyl C-5), 123.0 (pyridyl C-3), 125.7(═C(SAr)(CH(CO₂CH₂CH₃)₂)), 129.3 (benzylic C-1), 130.2 (benzylicC-2/C-6), 143.4 (pyridyl C-4, C—Cl), 150.6 (pyridyl C-6), 153.3(═C(CH₂CH₃)(pyridyl)), 158.4 (benzylic C-4, C—OCH₃), 159.1 (pyridylC-1), 167.2 (CH(CO₂CH₂CH₃)₂). LCMS m/z calcd for C₂₄H₂₈ClNO₅S 477([M+]). found 478 ([M+H]+).

Step 4. Ethyl8-chloro-1-ethyl-2-(4-methoxybenzylsulfanyl)-4-oxo-4H-quinolizine-3-carboxylate(4)

A solution of 3 (1.93 g, 4.04 mmol) in dimethyl sulfoxide (10.0 mL) isheated at 120° C. for 5.5 h, cooled to room temperature, andconcentrated under reduced pressure to give a yellowish brown oil thatsolidifies upon standing. The crude product is purified by flash columnchromatography (eluting with 20% ethyl acetate in methylene chloride; Rf0.53) to give pure 4 as a bright yellow solid. Because 4 oxidizes slowlyin solution under ambient conditions, the purified material is storedfor prolonged periods as a solid under an atmosphere of argon in theabsence of light.

mp 159-160° C. ¹H NMR (CDCl₃): δ 1.09 (t, J=7.5 Hz, 3H, CH₂CH₃), 1.44(t, J=7.0 Hz, 3H, CO₂CH₂CH₃), 2.87 (q, J=7.5 Hz, 2H, CH₂CH₃), 3.77 (s,3H, OCH₃), 4.10 (s, 2H, SCH₂), 4.51 (q, J=7.0 Hz, 2H, CO₂CH₂CH₃), 6.78(m, 2H, benzylic H-3/H-5), 7.01 (dd, J=8.0 Hz, 2.0 Hz, 1H, pyridoneH-7), 7.16 (m, 2H, benzylic H-2/H-6), 7.63 (d, J=2.0 Hz, 1H, pyridoneH-9), 9.16 (d, J=8.0 Hz, 1H, pyridone H-6). ¹³C{¹H} NMR (CDCl₃): δ 14.2(CO₂CH₂CH₃), 14.9 (CH₂CH₃), 22.0 (CH₂CH₃), 41.6 (SCH₂), 55.3 (OCH₃),62.0 (CO₂CH₂CH₃), 113.9 (benzylic C-3/C-5), 116.9 (C-7, CH), 119.1(C-1), 121.0 (C-3), 121.1 (C-9, CH), 128.8 (benzylic C-1), 130.0 (C-6,CH), 130.2 (benzylic C-2/C-6), 138.3 (C-8, C—Cl), 139.2 (C-9a), 144.5(C-2, C—SCH₂), 154.0 (C-4, NC(O)), 159.1 (benzylic C-4), 166.5(CO₂CH₂CH₃). LCMS m/z calcd for C₂₂H₂₂ClNO₄S 431 ([M+]). found 432([M+H]+). HRMS m/z calcd for C₂₂H₂₂ClNNaO₄S 454.0856 ([M+Na]+). found454.0853. Anal. Calcd for C₂₂H₂₂ClNO₄S: C, 61.18; H, 5.13; N, 3.24.Found: C, 61.33; H, 5.15; N, 3.33.

Step 5. Ethyl8-bromo-1-ethyl-2-mercapto-4-oxo-4H-quinolizine-3-carboxylate (5)

A solution containing 4 (1.0306 g, 2.39 mmol), trifluoroacetic acid (100g), and anhydrous anisole (20.0 mL) is heated at 40° C. for 23 h. Theresulting amber solution is evaporated under reduced pressure (˜1 mm Hg,40° C.) to give an oily residue containing ethyl8-bromo-1-ethyl-2-mercapto-4-oxo-4H-quinolizine-3-carboxylate (5) thatis used directly in the next step of the synthesis.

¹H NMR (CDCl₃): δ 1.22 (br, 3H, CH₂CH₃), 1.44 (t, J=7.0 Hz, 3H,CO₂CH₂CH₃), 2.88 (br, 2H, CH₂CH₃), 4.48 (br, 2H, CO₂CH₂CH₃), 5.85 (br,1H, SH), 6.92 (dd, J=8.0 Hz, 2.0 Hz, 1H, H-7), 7.53 (br, 1H, H-9), 9.12(d, J=8.0 Hz, 1H, H-6). LCMS m/z calcd for C₁₄H₁₄ClNO₃S 311 ([M+]).found 312 ([M+H]+).

Step 6.7-Chloro-9-ethyl-1-thia-2,4a-diazacyclopenta[b]naphthalene-3,4-dione (6)

A solution of sodium bicarbonate (2.00 g, 23.8 mmol) in water (60 mL) isadded to a solution of 5 (from above, ˜2.39 mmol) in tetrahydrofuran (60mL) at room temperature. Hydroxylamine-O-sulfonic acid (1.08 g, 9.55mmol) is added to this mixture as a solid and in one portion. Theresulting yellow mixture is stirred at room temperature for 3.5 h andquenched by addition of an aqueous solution of 5% hydrochloric acid (500mL). The precipitate that forms is collected by filtration, washed withdistilled water (3×50 mL), washed exhaustively with diethyl ether, anddried in vacuo to give pure 6 as a yellow solid. Because 6 decomposesslowly in solution under ambient conditions, the purified material isstored for prolonged periods as a solid under an atmosphere of argon inthe absence of light.

Rf 0.25 (methylene chloride/methanol/acetic acid≈200/10/1). mp 222-223°C. ¹H NMR (DMSO-d6): δ 1.14 (t, J=7.5 Hz, 3H, CH₂CH₃), 2.75 (q, J=7.5Hz, 2H, CH₂CH₃), 7.05 (dd, J=8.0 Hz, J=2.0 Hz, 1H, pyridone H-6), 7.87(d, J=2.0 Hz, 1H, pyridone H-8), 8.89 (d, J=8.0 Hz, 1H, pyridone H-5).¹³C{¹H} NMR (DMSO-d6): δ 13.3 (CH₂CH₃), 21.6 (CH₂CH₃), 104.0 (br, C-3a),106.1 (C-9), 114.4 (C-6, CH), 119.3 (C-8, CH), 129.7 (C-5, CH), 137.2(C-7, C—Cl), 137.5 (C-8a), 152.9 (C-4, NC(O)), 160.9 (br, C-9a, C—S),166.8 (C-3, CONS). LCMS m/z calcd for C₁₂H₉ClN₂O₂S 280 ([M+]). found 281([M+H]+). HRMS m/z calcd for C₁₂H₉ClN₂NaO₂S 302.9971 ([M+Na]+). found302.9969. Anal. Calc'd for C₁₂H₉ClN₂O₂S.H₂O: C, 48.24; H, 3.71; N, 9.38.Found: C, 48.06; H, 3.40; N, 9.16.

Step 6a.[2-¹⁵N]-7-Chloro-9-ethyl-1-thia-2,4a-diazacyclopenta[b]naphthalene-3,4-dione([2-¹⁵N]-7)

This compound is prepared as outlined above for 6 using[¹⁵N]hydroxylamine-O-sulfonic acid (98+ atom %). The ¹³C NMR spectrum of[2-¹⁵N]-6 is identical to that reported above for 6 with one exceptionthat follows. ¹³C{¹H} (DMSO-d6): δ 166.8 (d, 1JN—C=3.5 Hz). Becauseexchange of the N—H group of [2-¹⁵N]-6 is rapid (the corresponding broad1H resonance showed rapid H-D exchange with D₂O at room temperature), itis necessary to cool samples to obtain ¹⁵N NMR spectroscopic data. ¹⁵NNMR (DMF-d7, −50° C.): δ 104.6 (s). In accord with the ¹⁵N data recordedat low temperature, no 1JN-H coupling was observed in 1H NMR spectra of[2-¹⁵N]-7 recorded in DMF-d7 at temperatures ranging from +60 to −55° C.LCMS m/z calcd for C₁₂H₉ClN₁₅NO₂S ([M]+) 281. found 282 ([M+H]+).

Step 7. 10-ethyl-8-phenyl-2H-isothiazolo[5,4-b]quinolizine-3,4-dione (7)

Pd(PPh₃)₄ (3.1 mg, 6% mole), boronic acid or ester (2 equiv., 0.088mmole) and NaHCO₃ solution (1M, 0.2 mL, 4.5 equiv.) under argon at roomtemperature is added to a stirred solution of7-Bromo-9-ethyl-1-thia-2,4a-diazacyclopenta[b]naphthalene-3,4-dione0.044 mmole) in DMF (1 mL). The reaction mixture is degassed by bubblingargon through for 10 minutes at room temperature. The reaction tube issealed and then heated in a microwave (100 W, 130° C.) until thereaction reaches completion (10-20 minutes). The reaction mixture iscooled to room temperature and then filtered. The filtrate isconcentrated in vacuo. The residue is dissolved in a mixture ofDMF:CHCl₃:MeOH (0.5:3:0.5) (4 mL) and precipitated with diethyl ether.This dissolution and precipitation step is repeated 5 times. Theresulting light yellow solid is washed with water (3 mL) and dried toafford the title compound.

General information concerning the preparation of C—C bond at C-7 of apyridone core via Suzuki cross-coupling may be found in Otten, P. A. etal,. Bioconjugate Chem. (2001) 12: 203-212.

Example 2 Preparative HPLC Conditions

Preparative HPLC was performed using a YMC Pack Pro C18 150×20.0 mm 5 μmcolumn with an isocratic elution of 0.35 min at 90:10 water:acetonitrilecontaining 0.1% trifluoroacetic acid followed by a 23.6-min lineargradient elution from 90:10 to 10:90 at a flow rate of 18.9 mL/min withUV detection at 254 nm. Analytical HPLC was performed using a YMC PackPro C18 50×4.6 mm 5 μm column with an isocratic elution of 0.24 min at95:5 water:acetonitrile containing 0.1% trifluoroacetic acid followed bya 20-min linear gradient elution from 95:5 to 5:95 at a flow rate of 1.0mL/min with UV detection at 254 nm.

Example 39-Ethyl-7-pyridin-4-yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dionehydrochloride (8)

Tetrakis(triphenylphosphine)palladium(0) (26.1 mg, 0.023 mmol),4-pyridinylboronic acid (89.4 mg, 0.727 mmol), and a 1 M aqueoussolution of sodium bicarbonate (2.2 mL, 2.2 mmol) are added sequentiallyunder an atmosphere of argon, to a solution containing7-chloro-9-ethyl-1-thia-2,4a-diazacyclopenta[b]naphthalene-3,4-dione(40.9 mg, 0.145 mmol) and dimethylformamide (4.0 mL). The resultingorange mixture is irradiated in a microwave for 15 min (110° C.; 300 W)to give a red mixture. The solvent is evaporated under reduced pressure(˜10 mm Hg, 50° C.), and the remaining solid washed with diethyl ether(3×15 mL). The product is extracted from this solid using hot dimethylsulfoxide (5×2 mL) and purified by preparative HPLC. The purifiedproduct is isolated as the trifluoroacetate salt and converted to thehydrochloride salt by addition of a solution of hydrogen chloride (˜1.25M in methanol) followed by evaporation to give a deep red solid; thisprocess is repeated twice.

Purity by HPLC: 98.3%; tR=4.98 min. mp 289-290° C. dec. ¹H NMR(DMSO-d6): δ 1.22 (t, J=7.5 Hz, 3H, CH₂CH₃), 2.97 (q, J=7.5 Hz, 2H,CH₂CH₃), 7.52 (dd, J=8.0 Hz, J=1.5 Hz, 1H, pyridone H-6), 8.25 (d, J=1.5Hz, 1H, pyridone H-8), 8.40 (d, J=6.0 Hz, AA′ part of AA′XX′ spinsystem, 2H, pyridinyl H-3/H-5), 8.96 (d, J=6.0 Hz, XX′ part of AA′XX′spin system, 2H, pyridinyl H-2/H-6), 9.02 (d, J=8.0 Hz, 1H, pyridoneH-5). ¹³C{1H} NMR (DMSO-d6): δ 13.7 (CH₂CH₃), 21.8 (CH₂CH₃), 105.3(C-3a), 109.5 (C-9), 110.8 (C-6, CH), 121.4 (C-8, CH), 123.9 (pyridinylC-3/C-5), 128.8 (C-5, CH), 136.5 (C-7), 136.7 (C-8a), 143.7 (pyridinylC-2/C-6), 149.9 (pyridinyl C-4), 152.9 (C-4, NC(O)), 160.9 (C-9a, C—S),166.8 (C-3, CONS). LCMS m/z calcd for C₁₇H₁₃N₃O₂S 323 ([M+]+). found 324([M+H]+). HRMS m/z calcd for C₁₇H₁₄N₃O₂S 324.0807 ([M+H]+). found324.0807. Anal. Calcd for C₁₇H₁₃N₃O₂S.HCl.2.2H₂O: C, 51.11; H, 4.64; N,10.52. Found: C, 51.03; H, 4.54; N, 10.67. Analytical HPLC: tR=4.98 min.

Example 47-(2,6-Dimethylpyridin-4-yl)-9-ethyl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dionehydrochloride (9)

Tetrakis(triphenylphosphine)palladium(0) (25.9 mg, 0.022 mmol),2,6-dimethyl-4-pyridinylboronic acid (112.8 mg, 0.747 mmol), and a 1 Maqueous solution of sodium bicarbonate (2.3 mL, 2.3 mmol) are addedsequentially under an atmosphere of argon, to a solution containing7-chloro-9-ethyl-1-thia-2,4a-diazacyclopenta[b]naphthalene-3,4-dione(42.0 mg, 0.150 mmol) and dimethylformamide (4.0 mL). The resultingorange mixture is irradiated in a microwave for 15 min (110° C.; 300 W)to give a red mixture. The solvent is evaporated under reduced pressure(˜10 mm Hg, 50° C.), and the remaining solid washed with diethyl ether(3×15 mL). The product is extracted from this solid using hot dimethylsulfoxide (5×2 mL) and purified by preparative HPLC. The purifiedproduct is isolated as the trifluoroacetate salt and converted to thehydrochloride salt by addition of a solution of hydrogen chloride (˜1.25M in methanol) followed by evaporation to give a deep red solid; thisprocess is repeated twice.

Purity by HPLC: 99.6%; tR=5.80 min. mp 279-280° C. dec. ¹H NMR(DMSO-d6): δ 1.21 (t, J=7.5 Hz, 3H, CH₂CH₃), 2.80 (s, 6H, pyridinylCH₃), 2.96 (q, J=7.5 Hz, 2H, CH₂CH₃), 7.45 (dd, J=8.0 Hz, J=1.5 Hz, 1H,pyridone H-6), 8.20 (br s, 1H, pyridone H-8), 8.28 (s, 2H, pyridinylH-3/H-5), 8.96 (d, J=8.0 Hz, 1H, pyridone H-5). LCMS m/z calcd forCl₁₉H₁₇N₃O₂S 351 ([M+]). found 352 ([M+H]+). HRMS m/z calcd forC19H18N₃O₂S 352.1120 ([M+H]+). found 352.1120.

Example 57-(4-Aminomethylphenyl)-9-ethyl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione(10)

Under an atmosphere of argon, tetrakis(triphenylphosphine)palladium(0)(20.4 mg, 0.018 mmol), 4-aminomethylphenylboronic acid (110.0 mg, 0.587mmol), and a 1 M aqueous solution of sodium bicarbonate (1.75 mL, 1.75mmol) were added sequentially to a solution containing7-chloro-9-ethyl-1-thia-2,4a-diazacyclopenta[b]naphthalene-3,4-dione(33.0 mg, 0.118 mmol) and dimethylformamide (3.0 mL). The resultingorange mixture was irradiated in a microwave for 15 min (110° C.; 300 W)to give a red mixture. The solvent is evaporated under reduced pressure(˜10 mm Hg, 50° C.), and the remaining orange solid is washed withdiethyl ether (3×15 mL).

LCMS (APCI) m/z calcd for Cl₁₉H₁₇N₃O₂S 351 ([M+]). found 352 ([M+H]+,12%), 335 (100%).

¹H NMR (acetic acid-d4): δ 1.32 (t, J=7.5 Hz, 3H, CH₂CH₃), 2.90 (q,J=7.5 Hz, 2H, CH₂CH₃), 4.36 (s, 2H, CH₂N), 7.49 (dd, J=8.0 Hz, J=1.5 Hz,1H, pyridone H-6), 7.69 (d, J=8.5 Hz, AA′ part of AA′XX′ spin system,2H, phenyl H-3 and H-5), 7.91 (d, J=8.5 Hz, XX′ part of AA′XX′ spinsystem, 2H, phenyl H-2 and H-6), 7.94 (d, J=1.5 Hz, 1H, pyridone H-8),9.24 (d, J=8.0 Hz, 1H, pyridone H-5).

Analytical HPLC: tR=6.86 min.

Example 6 Preparation of1-Thia-2,4a,8-triaza-cyclopenta[b]naphthalene-diones

Example 7 Additional1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-diones

Additional compounds of Formula I are prepared by the methodsillustrated in Examples 1. Compounds of Formula I may also be preparedby the method disclosed by Otten, P. A. et al. (Bioconjugate Chem., 12:203-212 (2001) which is hereby incorporated by reference for itsteachings regarding the preparation of C—C bond at C-7 of the pyridonevia Suzuki cross-coupling.

Additional compounds of Formula I are listed in Table I.

TABLE 1 Cpd. # Structure Name 11

9-Cyclopropyl-6-fluoro-7-pyridin-4- yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 12

9-Cyclopropyl-6-fluoro-7-pyridin-3- yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 13

9-Cyclopropyl-6-fluoro-7-(6-methyl- pyridin-3-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 14

9-Cyclopropyl-6-fluoro-7-(1,2,3,4- tetrahydro-isoquinolin-6-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene- 3,4-dione 15

9-Cyclopropyl-7-(2,6-dimethyl- pyridin-4-yl)-6-fluoro-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4- dione 16

9-Cyclopropyl-6-fluoro-7-(1-methyl-2,3-dihydro-1H-isoindol-5-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene- 3,4-dione 17

9-Cyclopropyl-6-fluoro-7-pyrimidin-5- yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 18

3-(9-Cyclopropyl-6-fluoro-3,4-dioxo- 2,3-dihydro-4H-1-thia-2,4a-diaza-cyclopenta[b]naphthalen-7-yl)- banzonitrile 19

9-Cyclopropyl-6-fluoro-7-(1H-indol-2- yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 20

9-Cyclopropyl-6-fluoro-7-(4-hydroxy-3-methoxy-phenyl)-1-thia-2,4a-diaza- cyclopenta[b]naphthalene-3,4-dione21

7-(3-Amino-4-fluoro-phenyl)-9- cyclopropyl-6-fluoro-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4- dione 22

9-Cyclopropyl-6-fluoro-7-quinolin-6- yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 23

9-Cyclopropyl-6-fluoro-7-[4-(2- piperazin-1-yl-ethoxy)-phenyl]-1-thia-2,4a-diaza-cyclopenta[b]naphthalene- 3,4-dione 24

7-(4-Aminomethyl-phenyl)-9- cyclopropyl-6-fluoro-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4- dione 25

9-Cyclopropyl-6-fluoro-7-pyridin-3- tlethynyl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 26

4-[7-(4-Aminomethyl-phenyl)-9- cyclopropyl-6-fluoro-3,4-dioxo-3H,4H-1-thia-2,4a-diaza- cyclopenta[b]naphthalene-2-yl]-butyric acid 27

Acetic acid 9-cyclopropyl-6-fluoro-7-(6-methyl-pyridin-3-yl)-4-oxo-4H-1- thia-2,4a-diaza-cyclopenta[b]naphthalen-3-yl ester 28

5-Amino-9-cyclopropyl-6-fluoro-7- pyrimidin-5-yl-1-thia-2,4a-diaza-ccyclopenta[b]naphthalene-3,4-dione 29

6-Amino-9-cyclopropyl-7-pyridin-3- yl-1-thia-2,4a-diaza-ccyclopenta[b]naphthalene-3,4-dione 30

9-Cyclopropyl-6-fluoro-8-methoxy-7- (1-methyl-2,3-dihydro-1H-isoindol-5-yl)-1-thia-2,4a-diaza- cyclopenta[b]naphthalene-3,4-dioned 31

9-Cyclopropyl-8-methoxy-7-(1- methyl-2,3-dihydro-1H-isoindol-5-yl)-1-thia-2,4a-diaza- cyclopenta[b]naphthalene-3,4-dione 32

3-[6-Fluoro-9-(2-fluoro-cyclopropyl)-3,4-dioxo-2,3-dihydro-4H-1-thia-2,4a-diaza-cyclopenta[b]naphthalen-7-yl]- benzonitrile 33

9-(2,4-Difluoro-phenyl)-6-fluoro-7-(1,2,3,4-tetrahydro-issoquinolin-6-yl)- 1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione 34

9-Cyclopropyl-6-fluoro-7-(1H-indol-2- yl)-1-thia-2,4a,8-triaza-cyclopenta[b]naphthalene-3,4-dione 35

9-Cyclopropyl-6-fluoro-7-(4-hydroxy- 3-methoxy-phenyl)-1-thia-2,4a,8-triaza-cyclopenta[b]naphthalene-3,4- dione 36

7-(3-Amino-4-fluoro-phenyl)-9- ccyclopropyl-6-fluoro-1-thia-2,4a,8-triaza-cyclopenta[b]naphthalene-3,4- dione

Additionally the invention includes the following compounds of FormulaA, B, C, and D:

Formula A

Formula B

Formula C

Formula D

The arrays provided below disclose compounds of the general Formula A,B, C, and D in the above table.

The values “R₅” in these compounds are found in Array R5.

The values “R₆” in these compounds are found in Array R6.

The values “R₇” in these compounds are found in Array R7.

The values “R₈” in these compounds are found in Array R8.

The values “R₉” in these compounds are found in Array R9.

Each combination of 1 element from each of Array R5, Array R6, Array R7,Array R8, and Array R9 specifically discloses a discrete compound of theinvention.

For example [R₅-1][R₆-2][R₇-33][R₉-19] of Formula A is

9-Cyclopropyl-6-fluoro-7-pyridin-4-yl-1-thia-2,4a,8-diaza-cyclopenta[b]naphthalene-3,4-dione

ARRAY R₅ *—H *—F *—Cl R₅-1 R₅-2 R₅-3 *—Br *—I *—OH R₅-4 R₅-5 R₅-6 *—CN*—NH₂ *—NO₂ R₅-7 R₅-8 R₅-9 *—NHNH₂ *—CH₃ *—CH₂CH₃ R₅-10 R₅-11 R₅-12*—CH₂CH₂CH₃ *—CH₂CH₂CH₂CH₃ *—OCH₃ R₅-13 R₅-14 R₅-15 *—OCH₂CH₃*—OCH₂CH₂CH₃ *—OCH₂CH₂CH₂CH₃ R₅-16 R₅-17 R₅-18 *—NHCH₃ *—NHCH₂CH₃*—NHCH₂CH₂CH₃ R₅-19 R₅-20 R₅-21 *—NHCH₂CH₂CH₂CH₃ *—N(CH₃)₂ *—N(CH₂CH₃)₂R₅-22 R₅-23 R₅-24 *—N(CH₂CH₂CH₃)₂ *—N(CH₂CH₂CH₂CH₃)₂ *—NHNHCH₃ R₅-25R₅-26 R₅-27 *—NHNHCH₂CH₃ *—NHNHCH₂CH₂CH₃ *—NHNHCH₂CH₂CH₂CH₃ R₅-28 R₅-29R₅-30 *—NHN(CH₃)₂ *—NHN(CH₂CH₃)₂ *—NHN(CH₂CH₂CH₃)₂ R₅-31 R₅-32 R₅-33*—NHN(CH₂CH₂CH₂CH₃)₂ R₅-34

ARRAY R₆ *—H *—F *—Cl R₆-1 R₆-2 R₆-3 *—Br *—I *—OH R₆-4 R₆-5 R₆-6 *—CN*—NH₂ *—CH₃ R₆-7 R₆-8 R₆-9 *—CH₂CH₃ *—CH₂CH₂CH₃ *—CH₂CH₂CH₂CH₃ R₆-10R₆-11 R₆-12 *—OCH₃ *—OCH₂CH₃ *—OCH₂CH₂CH₃ R₆-13 R₆-14 R₆-15*—OCH₂CH₂CH₂CH₃ *—NHCH₃ *—NHCH₂CH₃ R₆-16 R₆-17 R₆-18 *—NHCH₂CH₂CH₃*—NHCH₂CH₂CH₂CH₃ *—N(CH₃)₂ R₆-19 R₆-20 R₆-21 *—N(CH₂CH₃)₂*—N(CH₂CH₂CH₃)₂ *—N(CH₂CH₂CH₂CH₃)₂ R₆-22 R₆-23 R₆-24 *—CF₃ *—OCF₃ R₆-25R₆-26

ARRAY R₇ *—Cl R₇-1 *—Br R₇-2 *—I R₇-3 *—O(SO₂)CF₃ R₇-4 *—N₂BF₄ R₇-5*—CH₃ R₇-6 *—CH₂CH₃ R₇-7 *—CH₂CH₂CH₂ R₇-8 *—CH₂CH₂CH₂CH₃ R₇-9*—CH₂CH₂CH₂CH₂CH₃ R₇-10 *—CH₂CH₂CH₂CH₂CH₂CH₃ R₇-11 *—CH₂CH₂CH₂CH₂CH₂CH₃R₇-12 *—CH═CHCH₃ R₇-13 *—CH═CHCH₂CH₃ R₇-14 *—CH═CHCH₂CH₂CH₃ R₇-15*—(C═O)—CH₃ R₇-16 *—(C═O)—CH₂CH₃ R₇-17 *—(C═O)—CH₂CH₂CH₃ R₇-18*—(C═O)NH—CH₃ R₇-19 *—C(═O)NH—CH₂CH₃ R₇-20 *—(C═O)NH—CH₂CH₂CH₃ R₇-21*—C≡C—CH₃ R₇-22 *—C≡C—CH₂CH₃ R₇-23 *—C≡C—CH₂CH₂CH₃ R₇-24

R₇-25

R₇-26

R₇-27

R₇-28

R₇-29

R₇-30

R₇-31

R₇-32

R₇-33

R₇-34

R₇-35

R₇-36

R₇-37

R₇-38

R₇-39

R₇-40

R₇-41

R₇-42

R₇-43

R₇-44

R₇-45

R₇-46

R₇-47

ARRAY R₈ *—H *—F *—Cl R₈-1 R₈-2 R₈-3 *—Br *—I *—OH R₈-4 R₈-5 R₈-6 *—CN*—NH₂ *—NO₂ R₈-7 R₈-8 R₈-9 *—NHNH₂ *—CH₃ *—CH₂CH₃ R₈-10 R₈-11 R₈-12*—CH₂CH₂CH₃ *—CH₂CH₂CH₂CH₃ *—OCH₃ R₈-13 R₈-14 R₈-15 *—OCH₂CH₃*—OCH₂CH₂CH₃ *—OCH₂CH₂CH₂CH₃ R₈-16 R₈-17 R₈-18 *—NHCH₃ *—NHCH₂CH₃*—NHCH₂CH₂CH₃ R₈-19 R₈-20 R₈-21 *—NHCH₂CH₂CH₂CH₃ *—NHNHCH₃ *—NHNHCH₂CH₃R₈-22 R₈-23 R₈-24 *—NHNHCH₂CH₂CH₃ *—NHNHCH₂CH₂CH₂CH₃ *—NHN(CH₃)₂ R₈-25R₈-26 R₈-27 *—NHN(CH₂CH₃)₂ *—NHN(CH₂CH₂CH₃)₂ *—NHN(CH₂CH₂CH₂CH₃)₂ R₈-28R₈-29 R₈-30 *—CF₃ *—OCF₃ R₈-31 R₈-32

ARRAY R₉ *—CH₃ R₉-I *—CH₂CH₃ R₉-2 *—CH₂CH₂CH₃ R₉-33 *—CH₂CH₂CH₂CH₃ R₉-4*—(CH₂)₄CH₃ R₉-5 *—(CH₂)₅CH₃ R₉-6 *—(CH₂)₆CH₃ R₉-7 *—(CH₂)₇CH₃ R₉-8*—OCH₃ R₈-9 *—OCH₂CH₃ R₉-10 *—OCH₂CH₂CH₃ R₉-11 *—OCH₂CH₂CH₂CH₃ R₉-12*—NHCH₃ R₉-13 *—NHCH₂CH₃ R₉-14 *—NHCH₂CH₂CH₃ R₉-15 *—NHCH₂CH₂CH₂CH₃R₉-16 *—NH(CH₃)₂ R₉-`17 *—NH(CH₂CH₃)₂ R₉-18

R₉-19

R₉-20

R₉-21

R₉-22

R₉-23

R₉-24 *—CF₃ R₉-25 *—OCF₃ R₉-26

Example 8 X-ray Crystal Structure of7-Chloro-9-ethyl-1-thia-2,4a-diazacyclopenta[b]naphthalene-3,4-dione (6)

Data Collection

A yellow blade crystal of C₁₂H₉ClN₂O₂S having approximate dimensions of0.25×0.10×0.10 mm³ was mounted with epoxy cement on the tip of a fineglass fiber. All measurements were made on a Nonius KappaCCDdiffractometer with graphite monochromated Mo—Kα radiation.

Cell constants and an orientation matrix for data collectioncorresponded to a primitive monoclinic cell with dimensions:

-   -   a=7.0574(14) Å    -   b=9.950(2) Å    -   c=16.661(3) Å    -   V=1165.6(4) Å³    -   α=90°    -   β=94.96(3)°    -   γ=90°

For Z=4 and F.W.=280.72, the calculated density is 1.600 g/cm³. Based ona statistical analysis of intensity distribution, and the successfulsolution and refinement of the structure, the space group was determinedto be P2₁/c (#14).

The data were collected at a temperature of 173(2) K to a maximum 2θvalue of 56.54°. Five omega scans consisting of 37, 29, 34, 14, and 17data frames, respectively, were collected with a frame width of 2.0° anda detector-to-crystal distance, Dx, of 35.0 mm. Each frame was exposedtwice (for the purpose of de-zingering) for a total of 40s. The dataframes were processed and scaled using the DENZO software package.

Data Reduction

A total of 4725 reflections were collected of which 2872 were unique andobserved (R_(int)=0.0321). The linear absorption coefficient, μ, forMo—Kα radiation is 5.00 cm⁻¹, and no absorption correction was applied.The data were corrected for Lorentz and polarization effects.

Structure Solution and Refinement

The structure was solved by direct methods and expanded using Fouriertechniques. The non-hydrogen atoms were refined anisotropically, andhydrogen atoms, with exceptions noted, were treated as idealizedcontributions. The final cycle of full-matrix least-squares refinement³on F was based on 2872 observed reflections (I>2.00σ(I)) and 167variable parameters and converged with unweighted and weighted agreementfactors of:R=Σ∥Fo|−|Fc∥/Σ|Fo|=0.0406R _(w) ={Σ[w(F _(o) ² −F _(c) ²)² ]/Σ[w(F _(o) ²)²]}^(1/2)=0.1030

The maximum and minimum peaks on the final difference Fourier mapcorresponded to 0.324 and −0.347 e⁻/Å³ respectively.

Example 9 Antimicrobial Activity of Compounds of the Invention

The antimicrobial activity of the compounds of the invention may beevaluated by a number of methods, including the following visual minimuminhibitory concentration (MIC) assay. This assay determines the minimumconcentration of compound required to inhibit growth of a bacterialstrain.

Minimum Inhibitory Concentration (MIC) Assay

Whole-cell antibacterial activity is determined by broth microdilutionusing conditions recommended by the National Committee for ClinicalLaboratory Standards (NCCLS). The minimum inhibitory concentration (MIC)was defined as the lowest concentration of antimicrobial agent tocompletely inhibit visible growth after 18-24 h at 37° C.

Test compounds are dissolved in DMSO and diluted 1:50 in Mueller-HintonII broth (Becton-Dickinson) to produce a 256 μg/ml stock solution. In a96-well microtiter plate, the compound solution is serially two-folddiluted in Mueller-Hinton II broth. After the compounds are diluted, a50 μg aliquot of the test organism (˜1×10⁶ cfu/mL) is added to each wellof the microtiter plate. The final test concentrations range from0.125-128 μg/mL. Inoculated plates are incubated in ambient air at 37°C. for 18 to 24 hours. The organisms selected for testing includedlaboratory strains S. aureus ATCC 29213 and E. coli ATCC 25922 (strainspurchased from American Type Culture Collection, Manassas, Va.). Theminimum inhibitory concentration (MIC) was determined as the lowestconcentration of compound that inhibited visible growth of the testorganism.

Topoisomerase IV Activity

Enzyme activity is measured by a decatenation assay that monitors theATP-dependent unlinking of DNA minicircles from kinetoplast DNA.Analysis is done by gel electrophoresis through 1% agarose/TBE gel. DNAis visualized with an Alpha Imager 2200 Analysis System and the IC₅₀determined by nonlinear regression analysis with Graphpad Prismsoftware. S. aureus topoismerase IV enzyme subunits, grlA and grlB, arepurified to homogeneity from over-expression plasmids in E. coli.

DNA Gyrase Activity

Enzyme activity is measured by a supercoiling assay that monitors theATP-dependent conversion of relaxed pBR₃₂₂ DNA to the supercoiled form.Analysis is done by gel electrophoresis through 1% agarose/TBE gel. DNAis visualized with an Alpha Imager 2200 Analysis System and the IC₅₀determined by nonlinear regression analysis with Graphpad Prismsoftware. E. coli DNA gyrase enzyme subunits, gyrA and gyrB, arepurified to homogeneity from over-expression constructs in E. coli.

Human Topoisomerase II Activity

Enzyme activity is measured by a DNA cleavage assay that monitorsgeneration of linear DNA from supercoiled pBR₃₂₂ DNA. Human p170topoisomerase II is purchased from TopoGen. Analysis is done byelectrophoresis through 1% agarose/TAE gel with 0.5 μg/mL ethidiumbromide, followed by visualization and quantification with Alpha Imager2200. The EC₂ value is defined as the effective concentration of drugrequired to enhance enzyme-mediated cleavage of double-stranded DNAtwofold.

Compounds 8 and 9 exhibit an MIC of 10 μg/ml against S. aureaus and E.coli when tested in the above assay for minimal inhibitory activity.These compounds also exhibit inhibition of e. coli DNA gyrasesupercoiling (IC₅₀<1 μM) and inhibition of S. aureus topoisomerase IVactivity (IC₅₀<10 μM). Compounds 8 and 9 do not inhibit S. aureus DNAgyrase supercoiling activity (IC₅₀>200 μM) and do not display activityagainst human topoisomerase II (EC2>150 μM). EC2 is the effectiveconcentration of drug required to enhance enzyme-mediated cleavage ofdouble-stranded DNA.

Example 10 Cell Viability Staining with Alamar Blue

To determine whether the microcidal effect observed against S. aureusand E. coli is specific to bacterial cells, compounds are screened forcell viability effects on several human cell types.

Optimal cell density is first determined by plating cells in a 96-wellplate standard sterile tissue culture plates in 100 μl media, 10% FBS atsix cell densities from 500 cells/well to 15,000 cells/well. A cell freewell containing only media is used as a control. Cells are incubated at37° C. in a 5% CO₂ incubator for 24 hours. 10% culture volume (10microliters) of Alamar Blue (Biosource, DAL 1100, 100 mL) is then added.Cells are incubated at 37° C. in a 5% CO₂ incubator and read in a VictorV plate reader, 544 nm excitation, 590 nm emission, at 3, 4, and 24hours after the addition of Alamar Blue. The cell number vs. change influorescence is plotted to determine linearity of signal vs. cellnumber. The optimal density varies between 500-15,000 cells/welldepending on the specific cell type. The optimal density is selectedbased on the highest number of cells that is still in the linearresponse range.

Determination of Compound Cytotoxicity

Cells are plated at optimal cell density in a standard sterile tissueculture 96 well plate, and incubated at 37° C. O/N in a 5% CO2incubator. 12 to 48 hours post-plating media is removed. The cells arewashed 1 or 2 times with 1×PBS and replaced with fresh media containingthe test compound in 1% DMSO. 24 to 72 hours after addition of compound,the media is removed, and the cells washed 1 to 2 times with 1×PBS.Fresh media containing 1/10 volume of Alamar Blue is then added. Platesare incubated 4 hours at 37° C. in a 5% CO2 incubator and read in aVictor V plate reader, 544 nm excitation, 590 nm emission.

Compounds are diluted to 20 micromolar in 1% DMSO and media and screenedin duplicate to obtain single concentration cytotoxicity data. Eightconcentration points from 0.78 micromolar to 100 micromolar, run induplicate, are used to determine cyctotoxicity CC50 values. Cells with1% DMSO and media are used as a negative control, compounds having aknown CC50 against a particular cell type are used as positive controls.

The change in fluorescence vs. concentration of test compound is plottedto determine the cytotoxicity of the compound.

Sample media conditions, optimal plating densities, and positive controlcompounds for two cell types screened are presented in Table II.

Preferred compounds disclosed in Example 1 and 3 to 6 exhibit CC50values greater than 10 uM against each of the cell lines listed below.Other cell types that may be used include but are not limited toBalb/3TC, CEM-SS, HeLa, HepG2, HT-29, MRC-5, SK-N—SH, U-87 MG, 293T, andHuh-7. More preferred are compounds with a CC₅₀ value greater than 50uM. Most preferred are compounds with a CC₅₀ value greater than 100 uM.

TABLE II CELL PLATING POSITIVE LINE MEDIA DENSITY CONTROL CHO 1. F-12Nutrient 7,000 cells/well Terfenadine (Chinese Mixture (Gibco CC₅₀ =hamster #11765-054) 4.3-6.5 μM ovary) containing 10% FBS, 1% Pen Strep,1.5 g/L Sodium Bicarbonate 2. McCoy's 5a medium, 10% FBS and PS/Gln HEP2 Minimum Essential 7,000 cells/well Terfenadine (laryngeal Medium -Alpha CC₅₀ = carcinoma) Medium (Gibco 3-5 μM # 12571-063) containing 10%FBS, 1% Pen Strep, 1.5 g/L Sodium Bicarbonate

Example 11 Pharmmaceutical Formulations

Examples 11A through 11D are examples of pharmaceutical compositionscontaining the compounds of Formula I. The abbreviation ‘A.M.’ standsfor an antimicrobial compound of the present invention.

Example 11A Capsules

20 grams of the A.M., 6 grams sodium lauryl sulfate, 56 grams starch, 56grams lactose, 0.8 grams colloidal silicon dioxide, and 1.2 gramsmagnesium stearate are vigorously stirred together. The resultingmixture is subsequently filled into 1000 suitable hardened gelatincapsules, comprising each 20 mg of the active ingredient.

Example 11B Film-Coated Tablets

Preparation of tablet core: A mixture of 10 grams of the A.M., 57 gramslactose and 20 grams starch is mixed well and thereafter humidified witha solution of 0.5 grams sodium dodecyl sulfate, and 1.0 gramspolyvinylpyrrolidone (KOLLIDON-K 90) in about 20 ml of water. The wetpowder mixture is sieved, dried, and sieved again. Then 100 gramsmicrocrystalline cellulose (AVICEL) and 15 grams hydrogenated vegetableoil (STEROTEX) are added. The whole is mixed well and compressed intotablets, giving 1000 tablets, each containing 10 mg of the activeingredient.

Coating: Ethyl cellulose (0.5 grams, ETHOCEL 22 CPS) in 15 ml ofdichloromethane is added to a solution of 1.0 grams methyl cellulose(Methocel 60 HG.RTM.) in 7.5 ml of denatured ethanol. Then 7.5 ml ofdichloromethane and 0.25 ml 1,2,3-propanetriol are added. Polyethyleneglycol (1.0 grams) is melted and dissolved in 7.5 ml of dichloromethaneand added to the cellulose-containing solution. Magnesium Octadecanoate(0.25 grams), 0.5 grams polyvinylpyrrolidone, and 3.0 ml of concentratedcolor suspension (OPASPRAY K-1-2109) are added and the whole mixturehomogenized. The tablet cores are coated with this mixture in a coatingapparatus.

Example 11C Injectible Solutions

(i) 1.8 grams methyl 4-hydroxybenzoate and 0.2 grams propyl4-hydroxybenzoate are dissolved in about 0.5 L of boiling water. Aftercooling to about 50° C., 4 grams lactic acid, 0.05 grams propyleneglycol, and 4 grams of the A.M are added while stirring. The solution iscooled to room temperature and supplemented with water for injectionq.s. giving a solution containing 4 mg/ml of A.M. The solution issterilized by filtration and filled in sterile containers.

(ii) 100.0 g of an acid salt of an A.M. of the invention is dissolved inboiling water. After cooling to about 50° C., 37.5 grams lactic acid(90% by weight) are added while stirring. The solution is cooled to roomtemperature and water is added to 1 L. The solution is sterilized byfiltration and filled in sterile containers.

(iii) 5.00 g of an acid salt of an A.M. of the invention is dissolved inboiling water. After cooling to about 50° C., 2.20 grams lactic acid(90% by weight) are added while stirring. The solution is cooled to roomtemperature and water is added to 100 ml.

Example 11D Cream

Phase I contains Sorbitan monostearate (2.0 g), Polyoxyethylene (20)sorbitan monostearate (1.5 g), Synthetic spermaceti (3.0 g) Cetylstearyl alcohol (10.0 g) and 2-Octyldodecanol (13.5 g). The phase Imixture is heated to 75° C., stirred and mixed. Phase II contains A.M.(1.0 g). Phase II is added to phase I, stirred and suspended. Phase IIIcontains Benzyl alcohol (1.0 g) and demineralized water (q.s. 100 g).Phase III is heated to 75° C. and added to phase II. The cream is mixedintensively and cooled slowly to room temperature, with furtherstirring. After cooling to room temperature the cream is homogenized.

1. A compound of Formula I

or a pharmaceutically acceptable salt thereof, wherein A₁ is S, O, SO, or SO₂; R₅ is hydrogen, halogen or amino R₆ is hydrogen, halogen, or amino, cyano; R₇ is XR_(A) where X is absent or —C≡C—, and R_(A) is phenyl, or a monocyclic or bicyclic 5- to 10- membered bicyclic saturated, partially unsaturated, or aromatic heterocylic group containing at least one nitrogen atom and bound via a carbon atom; each of which R_(A) is substituted with 0 to 5 substituents independently chosen from halogen, hydroxy, amino, cyano, C₁-C₆alkyl optionally substituted with amino, (C₁-C₆alkoxy)C₀-C₄alkyl, mono- and di-(C₁-C₄)alkylamino, ═NOR₁₀, where R₁₀ is C₁-C₄alkyl, and —OR_(D), where R_(D) is (C₂-C₆heterocycloalkyl)C₀-C₂alkyl; A₈ is CR₈; R₈ is hydrogen, C₁-C₄alkyl, C₁-C₄alkoxy; and R₉ is (C₃-C₇cycloalkyl)C₀-C₄alkyl or phenyl, each of which is substituted with 0 to 3 substituents independently chosen from halogen.
 2. A compound of Formula II

or a pharmaceutically acceptable salt thereof, wherein A₁ is S, O, SO, or SO₂; R₃ is hydrogen, C₁-C₆alkyl, C₁-C₆alkanoyl, mono- or di-C₁-C₆alkylcarbamate, or C₁-C₆alkylsulfonate; each of which is substituted with 0 to 3 substituents independently chosen from halogen, hydroxy, amino, cyano, nitro, C₁-C₄alkoxy, mono- and di-C₁-C₄alkylamino, C₁-C₂haloalkyl, and C₁-C₂haloalkoxy; R₅ is hydrogen, halogen or amino; R₆ is hydrogen, halogen, or amino; R₇ is XR_(A) where X is absent or —C≡C—, and R_(A) is phenyl or a monocyclic or bicyclic 5- to 10- membered bicyclic saturated, partially unsaturated, or aromatic heterocylic group containing at least one nitrogen atom and bound via a carbon atom; each of which R_(A) is substituted with 0 to 5 substituents independently chosen from halogen, hydroxy, amino, cyano, C₁-C₆alkyl optionally substituted with amino, (C₁-C₆alkoxy)C₀-C₄alkyl, mono- and di-(C₁-C₄)alkylamino, ═NOR₁₀, where R₁₀ is C₁-C₄alkyl, and —OR_(D), where R_(D) is (C₂-C₆heterocycloalkyl)C₀-C₂alkyl; R₈ is hydrogen, C₁-C₄alkyl, or C₁-C₄alkoxy; and R₉ is (C₃-C₇cycloalkyl)C₀-C₄alkyl, or phenyl, each of which is substituted with 0 to 3 substituents independently chosen from halogen.
 3. A compound or salt of claim 1, wherein A₁ is S.
 4. A compound or salt of claim 3 wherein R₅ is hydrogen.
 5. A compound or salt of claim 3 wherein: R₆ is hydrogen, fluoro, or amino.
 6. A compound or salt of claim 5 wherein R₆ is fluoro or hydrogen.
 7. A compound or salt of claim 3 wherein R₈ is hydrogen or methoxy.
 8. A compound or salt of claim 3 wherein R₉ is C₃-C₇cycloalkyl.
 9. A compound or salt of claim 8 wherein R₉ is cyclopropyl or 2,4-difluorophenyl.
 10. A compound or salt of claim 1 wherein R₇ is XR_(A) where X is absent or —C≡C—,and R_(A) is phenyl, dihydroindolyl, dihydroisoindolyl, dihydroquinolinyl, dihydroisoquinolinyl, indolyl, imidazolyl, isoindolyl, isoquinolinyl, isoxazolyl, piperazinyl, piperidinyl, pyrazinyl, pyridyl, pyrirnidinyl, pyrrolidinyl, pyrrolyl, pyrazolyl, quinolinyl, quinazolinyl, quinoxalinyl, thiazolyl, tetrahydropyridinyl, tetrahydroisoquionolinyl, or tetrahydroquinolinyl; group; each of which is substituted with 0 to 5 substituents independently chosen from halogen, hydroxy, amino, cyano, C₁-C₆alkyl optionally substituted with amino, (C₁-C₆alkoxy)C₀-C₄alkyl, mono- and di-(C₁-C₄)alkylamino, ═NOR₁₀, where R₁₀ is C₁-C₄alkyl, and —OR_(D), where R_(D) is C₂-C₆heterocycloalkyl)C₀-C₂alkyl.
 11. A compound or salt of claim 10 wherein R_(A) is a phenyl, dihydroindolyl, dihydroisoindolyl, dihydroquinolinyl, dihydroisoquinolinyl, indolyl, isoindolyl, isoquinolinyl, pyridyl, pyrirnidinyl, quinolinyl, tetrahydroisoquionolinyl, or tetrahydroquinolinyl; group; each of which is substituted with 0 to 5 substituents independently chosen from halogen, hydroxy, amino, cyano, C₁-C₄alkyl, C₁-C₄alkoxy, and mono- and di-(C₁-C₄)alkylamino.
 12. A compound or salt of claim 11 in which X is absent.
 13. A compound or salt of claim 12 in which R_(A) is phenyl or pyridyl, each of which is substituted with 0 to 3 substituents independently chosen from halogen, hydroxy, amino, cyano, C₁-C₄alkyl, C₁-C₄alkoxy, and mono- and di-(C₁-C₄)alkylamino.
 14. A pharmaceutical composition comprising a compound or salt of claim 1 together with a pharmaceutically acceptable carrier, diluent, or excipient.
 15. A compound or salt thereof of claim 1, wherein the compound is 9-Cyclopropyl-6-fluoro-7-pyridin-4-yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-pyridin-3-yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-(6-methyl-pyridin-3-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-(1,2,3,4-tetrahydro-isoquinolin-6-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-7-(2,6-dimethyl-pyridin-4-yl)-6-fluoro-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-(1-methyl-2,3-dihydro -1H-isoindol-5-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-pyrimidin-5-yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 3-(9-Cyclopropyl-6-fluoro-3,4-dioxo-2,3-dihydro-4H-1-thia-2,4a-diaza-cyclopenta[b]naphthalen-7-yl)-benzonitrile; 9-Cyclopropyl-6-fluoro-7-(1H-indol-2-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-(4-hydroxy-3-methoxy-phenyl)-1-thia-2,4a-diaza -cyclopenta[b]naphthalene-3,4-dione; 7-(3-Amino-4-fluoro-phenyl)-9-cyclopropyl-6-fluoro-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-quinolin-6-yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-[4-(2-piperazin-1-yl-ethoxy)-phenyl]-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; (missing cmp 25) 7-(4-Aminomethyl-phenyl)-9-cyclopropyl-6-fluoro-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-7-pyridin-3-ylethynyl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 4-[7-(4-Aminomethyl-phenyl)-9-cyclopropyl-6-fluoro-3,4-dioxo-3H,4H-1-thia-2,4a-diaza-cyclopenta[b]naphthalen-2-yl]-butyric acid; Acetic acid 9-cyclopropyl-6-fluoro-7-(6-methyl-pyridin-3-yl)-4-oxo-4H-1-thia-2,4a-diaza-cyclopenta[b]naphthalen-3-yl ester; 5-Amino-9-cyclopropyl-6-fluoro-7-pyrimidin-5-yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 6-Amino-9-cyclopropyl-7-pyridin-3-yl-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-6-fluoro-8-methoxy-7-(1-methyl-2,3-dihydro-1H-isoindol-5-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 9-Cyclopropyl-8-methoxy-7-(1-methyl-2,3-dihydro -1H-isoindol-5-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione; 3-[6-Fluoro-9-(2-fluoro-cyclopropyl)-3,4-dioxo-2,3-dihydro-4H-1-thia-2,4a-diaza-cyclopenta[b]naphthalen-7-yl]-benzonitrile; and 9-(2,4-Difluoro-phenyl)-6-fluoro-7-(1,2,3,4-tetrahydro-isoquinolin-6-yl)-1-thia-2,4a-diaza-cyclopenta[b]naphthalene-3,4-dione.
 16. A compound or salt of claim 13 in which R_(A) is a phenyl, pyridyl, pyrimidinyl, isoquinolinyl, quinolinyl, indolyl, pyrrolyl, isoindolyl, dihydroisoindolyl, tetrahydroisoquinolinyl, or tetrahydroisoquinolinyl group, each of which is substituted with 0 to 3 substituents independently chosen from halogen, hydroxy, amino, cyano, C₁-C₄alkyl, C₁-C₄alkoxy, mono- and di-(C1-C₄)alkylamino, C₁-C₂haloalkyl, and C₁-C₂haloalkoxy.
 17. A compound or salt of claim 16 in which R_(A) is phenyl, pyridyl-3-yl or pyrid-4-yl, each of which is substituted with 1 or 2 substituents independently chosen from fluoro, amino, hydroxy, cyano, and methyl.
 18. A compound or salt of claim 13 in which R_(A) is isoindol-5-yl, tetrahydroisoquinolin-5-yl, tetrahydroisoquinolin-6-yl, tetrahydroisoquinolin-7-yl, or tetrahydroisoquinolin-8-yl, each of which is substituted with 0 to 3 substituents independently chosen from C₁-C₃alkyl.
 19. A compound or salt of claim 10 in which R_(A) is dihydroindolyl, dihydroisoindolyl, tetrahydroisoquionolinyl, or tetrahydroquinolinyl, where R_(A) is substituted with 0 to 2 substituents independently chosen from halogen, hydroxy, amino, C₁-C₂alkyl, and C₁-C₂alkoxy. 